Comparing cell-free DNA patterns in dogs with sarcoma and healthy

Abstract

UNLABELLED: Plasma cell-free DNA (cfDNA) analysis holds potential to improve the detection of naturally occurring cancer in dogs, which can also serve as important model organisms for human cancer. However, there are limited comparative data on the characteristics of canine and human cfDNA. We characterized cfDNA fragmentation in 254 plasma samples from 54 healthy dogs and 54 dogs with naturally occurring sarcomas and compared them with 35 samples of human cfDNA. Using electrophoresis, whole-genome sequencing with both short and long reads, and multiplexed quantitative PCR, we assessed both fragment size distribution and fragment end sequences. We then trained a random forest classifier to distinguish healthy dogs from those with sarcomas based on cfDNA fragmentation features. Canine cfDNA fragment size distributions showed a striking deviation from humans, including a median of only 39% of fragments between 50 and 700 base pairs versus 84% in human samples. Fragment end nucleotides were more random in dogs than in humans at multiple size ranges. Similar to human patients with cancer, dogs with sarcomas had detectable copy-number changes and characteristically shorter cfDNA fragments relative to healthy dogs, enabling classification of cancer samples with 91% accuracy. Our results demonstrate key differences in cfDNA fragmentation between dogs and humans and highlight the potential for comparative translational research to advance blood-based early cancer detection in both species. SIGNIFICANCE: cfDNA analysis in dogs with naturally occurring cancers can enable precise detection of canine cancer and serve as a model system for human oncology. In this study, we observed differences in cfDNA fragmentation patterns between humans, healthy dogs, and dogs with sarcomas. Our results highlight the opportunity for comparative oncology research, the development of canine cfDNA diagnostics, and the investigation of biological mechanisms driving differences between humans and dogs.