Cancer detection in clinical practice and using blood-based liquid biopsy: A retrospective audit of over 350 dogs.

BACKGROUND: Guidelines-driven screening protocols for early cancer detection in dogs are lacking, and cancer often is detected at advanced stages. HYPOTHESIS/OBJECTIVES: To examine how cancer typically is detected in dogs and whether the addition of a next-generation sequencing-based "liquid biopsy" test to a wellness visit has the potential to enhance cancer detection. ANIMALS: Client-owned dogs with definitive cancer diagnoses enrolled in a clinical validation study for a novel blood-based multicancer early detection test. METHODS: Retrospective medical record review was performed to establish the history and presenting complaint that ultimately led to a definitive cancer diagnosis. Blood samples were subjected to DNA extraction, library preparation, and next-generation sequencing. Sequencing data were analyzed using an internally developed bioinformatics pipeline to detect genomic alterations associated with the presence of cancer. RESULTS: In an unselected cohort of 359 cancer-diagnosed dogs, 4% of cases were detected during a wellness visit, 8% were detected incidentally, and 88% were detected after the owner reported clinical signs suggestive of cancer. Liquid biopsy detected disease in 54.7% (95% confidence interval [CI], 49.5%-59.8%) of patients, including 32% of dogs with early-stage cancer, 48% of preclinical dogs, and 84% of dogs with advanced-stage disease. CONCLUSIONS/CLINICAL IMPORTANCE: Most cases of cancer were diagnosed after the onset of clinical signs; only 4% of dogs had cancer detected using the current standard of care (i.e., wellness visit). Liquid biopsy has the potential to increase detection of cancer when added to a dog's wellness visit.

Multi-institutional retrospective study of canine foot pad malignant melanomas: 20 cases.

Melanomas arising from the foot pad are a rare clinical entity in dogs. The biologic behaviour of foot pad malignant melanoma is not well understood, and these tumours are infrequently described. The objective of this study was to evaluate the clinical characteristics of primary canine foot pad melanoma in a larger cohort of patients. Eligible cases were solicited from the American College of Veterinary Internal Medicine (ACVIM) Oncology listserv for retrospective review. Included dogs had a cytologic and/or histologic diagnosis of foot pad melanoma evaluated by a board-certified clinical or anatomic pathologist. Dogs with cutaneous, oral, digital, subungual or interdigital melanomas were excluded. A total of 20 cases were included. Eleven dogs received various adjuvant therapies including chemotherapy, radiation therapy, and/or the ONCEPT canine melanoma vaccine following surgery. At diagnosis, regional lymph node metastasis was observed in four dogs (20%). Seven dogs developed subsequent regional and/or distant metastasis for an overall metastatic rate of 55%. The progression-free interval (PFI) was 101 days (range, 20-960 days). The median survival time (MST) was 240 days (range, 25-479 days). For dogs receiving adjuvant therapy, the MST was 159 days (range, 25-387 days). Canine foot pad melanoma is a rare neoplasm that can exhibit an aggressive behaviour.

Clinical validation of a next-generation sequencing-based multi-cancer early detection "liquid biopsy" blood test in over 1,000 dogs using an independent testing set: The CANcer Detection in Dogs (CANDiD) study.

Cancer is the leading cause of death in dogs, yet there are no established screening paradigms for early detection. Liquid biopsy methods that interrogate cancer-derived genomic alterations in cell-free DNA in blood are being adopted for multi-cancer early detection in human medicine and are now available for veterinary use. The CANcer Detection in Dogs (CANDiD) study is an international, multi-center clinical study designed to validate the performance of a novel multi-cancer early detection "liquid biopsy" test developed for noninvasive detection and characterization of cancer in dogs using next-generation sequencing (NGS) of blood-derived DNA; study results are reported here. In total, 1,358 cancer-diagnosed and presumably cancer-free dogs were enrolled in the study, representing the range of breeds, weights, ages, and cancer types seen in routine clinical practice; 1,100 subjects met inclusion criteria for analysis and were used in the validation of the test. Overall, the liquid biopsy test demonstrated a 54.7% (95% CI: 49.3-60.0%) sensitivity and a 98.5% (95% CI: 97.0-99.3%) specificity. For three of the most aggressive canine cancers (lymphoma, hemangiosarcoma, osteosarcoma), the detection rate was 85.4% (95% CI: 78.4-90.9%); and for eight of the most common canine cancers (lymphoma, hemangiosarcoma, osteosarcoma, soft tissue sarcoma, mast cell tumor, mammary gland carcinoma, anal sac adenocarcinoma, malignant melanoma), the detection rate was 61.9% (95% CI: 55.3-68.1%). The test detected cancer signal in patients representing 30 distinct cancer types and provided a Cancer Signal Origin prediction for a subset of patients with hematological malignancies. Furthermore, the test accurately detected cancer signal in four presumably cancer-free subjects before the onset of clinical signs, further supporting the utility of liquid biopsy as an early detection test. Taken together, these findings demonstrate that NGS-based liquid biopsy can offer a novel option for noninvasive multi-cancer detection in dogs.

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.

Presumptive increase in protein-bound serum calcium in a dog with multiple myeloma.

An 11-year-old male castrated Australian Shepherd was presented with a history of lethargy, panting, and weight loss for 1 month. Physical examination revealed a moderately enlarged spleen. Laboratory abnormalities included thrombocytopenia and marked hypercalcemia, with hyperglobulinemia, hypoalbuminemia, and a monoclonal spike in the beta-globulin region on serum protein electrophoresis. Serum total calcium concentration was markedly increased (16.5 mg/dL, reference interval 8.9-11.4 mg/dL) but ionized calcium concentration (1.39 mmol/L) was within the reference interval (1.25-1.45 mmol/L). Isosthenuria was noted, but the dog was not polyuric or polydipsic. Serum parathyroid hormone concentration was within reference limits and parathyroid hormone-related peptide concentration was 0 pmol/L. Radiographic findings were largely unremarkable. Results of cytologic evaluation of a fine-needle aspirate specimen from the spleen indicated plasma cell neoplasia. Based on the results of the electrophoresis, splenic aspirates, radiographs, and hypercalcemia, a diagnosis of splenic multiple myeloma was made. The marked hypercalcemia, normal ionized calcium and parathyroid hormone concentrations, and lack of osteolytic lesions indicated a presumptive increase in protein-bound serum calcium, likely due to binding to molecules of the paraprotein (M protein). Protein binding of calcium in dogs with multiple myeloma should be considered as a potential mechanism of elevated total serum calcium concentration.

Multidrug Analyses in Patients Distinguish Efficacious Cancer Agents Based on Both Tumor Cell Killing and Immunomodulation.

The vision of a precision medicine-guided approach to novel cancer drug development is challenged by high intratumor heterogeneity and interpatient diversity. This complexity is rarely modeled accurately during preclinical drug development, hampering predictions of clinical drug efficacy. To address this issue, we developed Comparative In Vivo Oncology (CIVO) arrayed microinjection technology to test tumor responsiveness to simultaneous microdoses of multiple drugs directly in a patient's tumor. Here, in a study of 18 canine patients with soft tissue sarcoma (STS), CIVO captured complex, patient-specific tumor responses encompassing both cancer cells and multiple immune infiltrates following localized exposure to different chemotherapy agents. CIVO also classified patient-specific tumor resistance to the most effective agent, doxorubicin, and further enabled assessment of a preclinical autophagy inhibitor, PS-1001, to reverse doxorubicin resistance. In a CIVO-identified subset of doxorubicin-resistant tumors, PS-1001 resulted in enhanced antitumor activity, increased infiltration of macrophages, and skewed this infiltrate toward M1 polarization. The ability to evaluate and cross-compare multiple drugs and drug combinations simultaneously in living tumors and across a diverse immunocompetent patient population may provide a foundation from which to make informed drug development decisions. This method also represents a viable functional approach to complement current precision oncology strategies. Cancer Res; 77(11); 2869-80. ©2017 AACR.