Multicancer detection in dogs with cell-free DNA

dvm360dvm360 January 2022
Volume 53
Issue 1
San Diego

Novel liquid biopsy uses blood samples to detect cancer-associated genomic alterations in dogs.

Early detection and treatment for cancer in dogs is critical for improving outcomes for these patients. Despite cancer being a leading cause of canine death1, Andi Flory, DVM, DACVIM (Oncology), said many patients do not receive a diagnosis until after clinical signs have developed. While presenting a session at the Fetch dvm360® Conference in San Diego, California, Flory said canine cancer is typically diagnosed because of the presence of clinical signs, at which point cancer is often at an advanced stage, making the prognosis poor and the case more challenging to manage.2

In her session—" Liquid Biopsy for Noninvasive Multi-cancer Detection in Dogs,” sponsored by PetDx—Flory discussed how improved patient outcomes may be achieved with early cancer identification and treatment for the disease with liquid biopsy testing. She noted that despite the need, cancer screening paradigms do not exist for dogs, like they do for humans.1,2

“Unfortunately, in veterinary medicine, cancer screening guidelines are lacking,” said Flory.2

Annual or biannual physical exams and routine minimum database testing such as chemistry and urinalysis are unable to detect many cancers in dogs, she added, but noninvasive liquid biopsies that focus on high-risk canine populations, including older dogs and predisposed breeds, may help identify cancer at an earlier stage.2

“Early detection has been shown to drive better clinical outcomes in humans such as increased life expectancy and higher rates of achieving cure or long-term control following curative-intent interventions. It stands to reason those similar benefits could be achieved in canine patients with the availability of early detection screening tools,” said Flory.2

Screening with cell-free DNA

As liquid biopsies are evolving, cell-free DNA (cfDNA) is currently the most promising blood-based biomarker for detecting canine cancer, according to Flory. Composed of DNA fragments that are released into circulation upon cell death by apoptosis or necrosis, cfDNA is released by noncancerous cells as well as tumor cells. cfDNA can be analyzed with next-generation sequencing (NGS) technology to detect the presence of cancer.1

NGS can detect multiple classes of genomic alterations associated with cancer. When found in cfDNA, these genomic alterations indicate the presence of tumor cells in a patient’s body1, according to Flory.

The OncoK9 multi-cancer early detection test by PetDx is a novel, blood-based liquid biopsy that finds cancer-associated genomic alterations. In the Fetch talk, Flory said the OncoK9 test has proven ability to detect more than 20 cancer types, including high detection rates in many of the most common canine cancers, and has a low false-positive rate.2

Flory also noted the OncoK9 test requires a simple, in-clinic blood draw that does not require fasting, and the sample does not need to be processed in the clinic nor does it require refrigeration.2

Efficacy of blood-based liquid biopsy

According to Flory, blood-based liquid biopsy testing is poised to change the way we detect, characterize, and monitor cancer in dogs. In the Fetch presentation, Flory cited a proof-of-concept study as demonstration of the test’s capability.1 In the published study, for which Flory is a co-author, 11 dogs with confirmed cancer diagnoses and 5 dogs presumed to be cancer-free provided blood samples for investigation. Nine of the 11 dogs with cancer also had tumor tissue samples collected during surgery.3

In 5 of the 9 studied dogs with cancer and matched tumor and plasma samples, presurgical liquid biopsy testing identified genomic alterations that matched alterations independently detected in corresponding tumor tissue samples, according to investigators. Additionally, the presurgical liquid biopsy test detected alterations found in same-subject spatially separated tissue samples. This observation shows the potential of blood-based testing for comprehensive genomic profiling of heterogeneous tumors, according to investigators. The investigators also found that among the 3 patients with postsurgical blood samples, genomic alterations remained detectable in 1 patient with incomplete tumor resection, which suggests there is utility for noninvasive detection of minimal residual disease after curative-intent treatment.3

Using liquid biopsy in cancer care

According to Flory, there are 6 clinical scenarios where blood-based liquid biopsy can be employed. They are as follows2:

  • Screening in asymptomatic high-risk patients
  • Aid in diagnosis for patients in which there is a clinical suspicion of cancer
  • Targeted treatment selection
  • Detection of minimal residual disease following surgery
  • Response monitoring during cancer therapy
  • Recurrence monitoring after cancer therapy has been completed

“Importantly, liquid biopsy has demonstrated clinical utility in all 6 use cases in humans, allowing cancer detection and characterization and cancer therapy monitoring,” said Flory.

According to Flory, postsurgical treatments to prolong cancer control such as radiation, chemotherapy, and/or immunotherapy, are often recommended for patients with cancers that have high rates of recurrence or metastasis. Liquid biopsy can aid in detecting cancer remaining in the body that may lead to relapse or recurrence. Conventional methods such as imaging and physical exam may prove more challenging for detecting minimal residual disease than a liquid biopsy relying on cfDNA.1,2


Despite its’ benefits, liquid biopsy is not a solution to every challenge in veterinary cancer management, according to Flory. Limitations to this technology exist, including low circulating tumor DNA shedding by certain cancers such as those seen in central nervous system tumors, smaller-sized tumors, and in early disease.1,2

“Additionally, there are challenges that must be addressed for the clinical use of liquid biopsy solutions. For example, defining the genomic regions known to have clinical relevance to cancer, including those orthologous to cancer-associated genomic regions in humans, is a major effort that requires significant time and expertise,” Flory noted. “In this new field in veterinary medicine, it will be extremely important that any candidate liquid biopsy solution undergo validation at a level like that expected for human use to maximize the benefit for veterinary patients and clinicians.”2

Furthermore, Flory suggested that clinical validation be performed in “adequately sized” cohorts of canine subjects, with a variety of cancers and with dogs that are presumably cancer-free. Flory said future research will likely aim to study the influence of liquid biopsy on case management in dogs and research is needed to examine patients diagnosed at an earlier time point when clinical outcomes are likely to be improved.2

Establishing whether earlier detection of minimal residual disease, recurrence, and response to treatment could provide earlier opportunities for intervention, leading to improved outcomes, is also important, according to Flory. And using this liquid biopsy as a routine screening tool may prove beneficial for creating custom treatment plans for patients.1,2


  1. Chibuk J, Flory A, Kruglyak KM, et al. Horizons in Veterinary Precision Oncology: Fundamentals of Cancer Genomics and Applications of Liquid Biopsy for the Detection, Characterization, and Management of Cancer in Dogs. Front Vet Sci. 2021;8:664718. doi:10.3389/fvets.2021.664718.
  2. Flory A. Liquid biopsies for noninvasive multicancer detection in dogs. Presented at: Fetch dvm360® Conference; San Diego, California. December 2-5, 2021
  3. Kruglyak KM, Chibuk J, McLennan L, et al. Blood-based liquid biopsy for comprehensive cancer genomic profiling using next-generation sequencing: an emerging paradigm for non-invasive cancer detection and management in dogs. Front Vet Sci. 2021;8:698. doi:10.3389/fvets.2021.704835.
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