Cell-Free DNA as a Diagnostic and Prognostic Biomarker in Dogs With Tumors

Cell-free DNA (cfDNA) is derived from apoptosis/necrosis, active cellular secretion, and lysis of circulating cancer cells or micrometastases. In humans, cfDNA is widely used in cancer diagnosis, but veterinary research has yet to be actively conducted to establish it as a cancer biomarker. This retrospective study analyzed cfDNA levels in samples collected from dogs with neoplastic disease (n = 38), clinically ill dogs without neoplasia (n = 47), and healthy dogs (n = 35). cfDNA levels and clinical data were compared among groups, and prognostic analyses were performed within the neoplastic group. Furthermore, continual cfDNA measurements were performed during the chemotherapy of six dogs with lymphoma. Dogs with neoplasia showed significantly higher cfDNA concentrations than dogs without neoplasm, and the cfDNA oncentration in the lymphoid neoplasia group was significantly elevated among all neoplastic groups. Dogs with neoplasia and a plasma cfDNA concentration above 1,247.5 μg/L had shorter survival rates than those with levels below this threshold (26.5 vs. 86.1%, respectively, P < 0.05). In cases with complete remission in response to chemotherapy, the cfDNA concentration was significantly decreased compared with the first visit, whereas the cfDNA concentration was increased in cases with disease progression or death. Interestingly, a significant correlation was found between lymph node diameter and cfDNA concentration in dogs with multicentric lymphoma (R2 = 0.26, P < 0.01). These data suggest that changes in cfDNA concentration could be used as a diagnostic biomarker for canine neoplasia. Furthermore, increased plasma DNA levels might be associated with shorter survival time, and cfDNA concentrations may reflect the response to chemotherapy.

L-Asparaginase, Doxorubicin, Vincristine, and Prednisolone (LHOP) Chemotherapy as a First-Line Treatment for Dogs with Multicentric Lymphoma

Cyclophosphamide exhibits the weakest therapeutic effect compared with vincristine and doxorubicin in the CHOP (C, cyclophosphamide; H, doxorubicin; O, vincristine; and P, prednisolone) chemotherapeutic protocol for the treatment of canine lymphoma. Twenty dogs with multicentric lymphoma were treated using the LHOP protocol, which used l-asparaginase in place of cyclophosphamide, and the outcomes were historically compared with those of dogs that received CHOP chemotherapy in the same institution. No significant differences were found in age (p = 0.107), body weight (p = 0.051), sex (p = 0.453), clinical stage V (p = 1), substage b (p = 0.573), T-cell phenotype (p = 0.340), overall response (p = 1), and hypercalcaemia status (p = 1) between the LHOP and CHOP groups. The adverse effects of l-asparaginase were well tolerated and self-limiting. The median PFS (progression-free survival) and median ST (survival time) in the LHOP group were 344 days (range: 28–940 days) and 344 days (range: 70–940 days), respectively. The median PFS and median ST in the CHOP group were 234 days (range: 49–1822 days) and 314 days (range: 50–1822 days), respectively. The dogs that received LHOP chemotherapy had a significantly longer PFS than the dogs that received CHOP chemotherapy (p = 0.001). No significant difference was observed in ST between the LHOP and CHOP groups (p = 0.131). Our study findings thus indicate that the LHOP protocol can be used as a first-line chemotherapeutic protocol in canine multicentric lymphoma.

Characterizing Circulating Nucleosomes in the Plasma of Dogs with Lymphoma

Background: Nucleosomes consist of DNA wrapped around a histone octamer core like beads on a string so that DNA can be condensed as chromatin into chromosomes. Diseases such as cancer or inflammation lead to cell death where chromatin is fragmentated and released as mononucleosomes into the blood. The Nu.QTM H3.1 assay measures total nucleosome concentration in plasma of humans and has been used to detect and identify cancer even at early stages. The objectives of this study were to determine if nucleosome levels could be used to distinguish between healthy dogs and dogs with various stages of lymphoma (LSA) using the Nu.Q™ H3.1 assay. A total of 126 dogs diagnosed with LSA and 134 healthy controls were recruited for this study. Plasma was collected from each dog and stored in K2-EDTA tubes. The LSA patient samples were recruited from TAMU or purchased from various biobanks. All control cases were recruited from TAMU. Samples were also collected longitudinally from 3 dogs undergoing treatment for multicentric lymphoma at TAMU as a pilot study to investigate the pattern of nucleosome concentrations in plasma during treatment. Results: Dogs with LSA had an approximately 7-fold increase in their plasma nucleosome concentrations compared to controls (AUC 87.8%). Nucleosome concentrations increased with cancer stage and dogs with B cell lymphomas had significantly higher nucleosome concentrations than dogs with T cell lymphomas. Nucleosome concentrations from serially monitored patients were elevated at diagnosis and progression with subsequent decreases in nucleosome concentration that corresponded to clinically detectable responses to therapy. Conclusions: The Nu.QTM H3.1 assay was able to reliably detect elevated nucleosome concentrations in the plasma of dogs with LSA. Furthermore, it appears that nucleosomes are useful for differentiating cancer from healthy individuals in canines. Results from serially monitored patients indicate that nucleosomes could be an objective monitoring tool for remission status in canine lymphoma patients.

Cardiomyopathy in a dog with multicentric lymphoma following treatment with several anthracyclines

Background: Canine lymphoma is one of the most frequently occurring malignant neoplasms in dogs. Anthracycline-based chemotherapy for the treatment of canine lymphoma is very effective; however, there is not enough evidence for the development of cardiac toxicity using several anthracyclines as chemotherapeutic agents. Case Description: An 8-year-old, castrated, mixed-breed dog was diagnosed with multicentric lymphoma and received multi-agent chemotherapy. Complete remission was achieved, but the patient had a relapse of lymphoma. After third-line chemotherapy with epirubicin, the patient was diagnosed with dilated cardiomyopathy. The total cumulative doses of doxorubicin, mitoxantrone, and epirubicin were 125, 8, and 125 mg/m2, respectively. Although the patient was treated with cardiac drugs and clinically stabilized, the patient had a relapse of lymphoma and died shortly after the diagnosis of cardiomyopathy. Conclusion: The patient was suspected to have anthracycline-induced cardiomyopathy. Further studies are required to establish prevention and management strategies for dogs receiving potentially cardiotoxic therapies, such as anthracyclines.