Elevated levels of IL-12/IL-23p40 in Nova Scotia Duck Tolling Retrievers with autoimmune disease and lymphoma.

The Nova Scotia Duck Tolling Retriever (NSDTR) is predisposed to immune mediated rheumatic disease (IMRD), steroid-responsive meningitis-arteritis (SRMA) and certain forms of cancer. Cytokines are the main regulators of the immune system. Interleukin 2 is a cytokine involved in activation of T regulatory cells, playing a role in central tolerance and tumor immunity. Interleukin 12 and interleukin 23 share the same subunit, p40, and are both pro-inflammatory cytokines. The aim of this study was to compare levels of IL-2 in healthy NSDTRs to those with cancer or autoimmune disease and to compare levels of IL-12/IL-23p40 in healthy NSDTRs and beagles versus NSDTRs with cancer or autoimmune disease. 62 dogs were included in the analysis of IL-12/IL-23p40; healthy NSDTRs (n = 16), healthy beagles (n = 16), NSDTRs autoimmune (n = 18) and NDSTRs lymphoma/mastocytoma (n = 12) and 68 dogs for IL-2; healthy (n = 20), autoimmune (n = 36) and lymphoma/mastocytoma/adenocarcinoma (n = 12). NSDTRs with autoimmune disease had higher levels of IL-12/IL-23p40 compared to healthy dogs (p = 0.008). NSDTRs with lymphoma also had higher levels of IL-12/IL-23p40 compared to healthy NSDTRs (p = 0.002). There was no difference in levels of IL-2 between healthy and diseased NSDTR. Statistical analysis was performed using Bonferroni corrections for multiple testing. These findings can contribute to the knowledge of autoimmune disease and cancer in dogs.

Stage Migration in Canine Multicentric Lymphoma: Impact of Diagnostic Techniques on Assessing Disease Extent.

BACKGROUND/AIM: Stage migration, a phenomenon triggered by technological advancements allowing more sensitive tumor spread detection, results in alterations in the distribution of cancer stages within a population. Canine multicentric lymphoma is staged I to V based on the affected anatomic site(s) and substage a or b depending on the presence of tumor-related clinical signs. The primary objective of this study was to assess the influence of various diagnostic techniques on staging accuracy and determine whether multiple staging methods lead to significant stage migration, impacting the reliability of disease stage assignments. MATERIALS AND METHODS: Dogs cytologically diagnosed with multicentric lymphoma were staged using four different staging methods (A-D): A (physical examination, hemogram, blood smear), B (A plus thoracic X-ray, abdominal ultrasound), C (B plus liver and spleen cytology) and D (C plus bone marrow cytology). RESULTS: Twenty-three dogs were enrolled: 16 females (70%) and seven males (30%). Regarding immunophenotype, 21 dogs (91.3%) were B-cell and two dogs (8.7%) were T-cell. Stage migration was observed between all staging methods. Between A and B, 12 animals migrated from stage III to stage IV. Between B and C, four animals migrated, three to a higher stage (stage III to IV) and one to a lower stage (stage IV to III). Between C and D, one animal migrated from stage IV to V. The differences between staging methods A and B were statistically significant (p≤0.001). CONCLUSION: Stage migration in canine multicentric lymphoma depends on the diagnostic methods used and reinforces the need to use standardized staging methods to avoid it.

Evaluation of clinical response and prognostic factors in canine multicentric lymphoma treated with first rescue therapy.

Despite an initial strong response in most dogs with multicentric lymphoma treated with chemotherapy, relapse remains common. There is no clearly superior first rescue protocol described either for resistant or relapsed canine multicentric lymphoma. The objectives of this study were to assess clinical response and outcomes for canine multicentric lymphoma treated with first rescue protocols. The secondary objective was to assess prognostic variables for dogs undergoing these protocols. This was a bi-institutional retrospective cohort study. Two hundred and sixty-five dogs were treated with first rescue chemotherapy, including anthracycline-based combination chemotherapy (CHOP-like, n = 50), nitrosourea alkylating agent-rich chemotherapy (n = 45), anthracycline-based or related compound chemotherapy (n = 34), or nitrosourea single-agent chemotherapy (n = 136). The overall median progression free survival time of first rescue protocol was 56.0 days (0-455 days). Important prognostic factors identified for first rescue protocol included the attainment of a complete response to the first rescue chemotherapy (p < .001), the use of a CHOP-like first rescue protocol (p = .009), duration of first remission (HR 0.997, p = .028), and if prednisolone was included in the first rescue protocol (HR 0.41, p = .003). Adverse events (AE) were common, with 81.1% of dogs experiencing at least one AE during first rescue chemotherapy. This study highlights the need for improved first rescue therapies to provide durable remission in canine resistant or relapsed lymphoma.

Alternating rabacfosadine and doxorubicin for treatment of naïve canine lymphoma.

The current standard of care treatment for canine lymphoma is a multi-agent, CHOP-based chemotherapy protocol. Single agent doxorubicin (DOX) is less burdensome; however, multi-agent chemotherapy protocols are often superior. The recently approved drug rabacfosadine (RAB, Tanovea) provides an attractive option for combination therapy with DOX, as both drugs demonstrate efficacy against lymphoma and possess different mechanisms of action. A previous study evaluating alternating RAB/DOX reported an overall response rate (ORR) of 84%, with a median progression-free survival time (PFS) of 194 days. The aim of this prospective trial was to evaluate the same protocol in an additional population of dogs. Fifty-nine dogs with treatment naïve lymphoma were enrolled. RAB (1.0 mg/kg IV) was alternated with DOX (30 mg/m2 IV) every 21 days for up to six total treatments (3 cycles). Response assessment and adverse event (AE) evaluation were performed every 21 days using VCOG criteria. The ORR was 93% (79% CR, 14% PR). The median time to maximal response was 21.5 days; median PFS was 199 days. T cell immunophenotype and lack of treatment response were predictive of inferior outcomes. AEs were mostly gastrointestinal. Six dogs developed presumed or confirmed pulmonary fibrosis; four were grade 5. One dog experienced grade 3 extravasation injury with RAB that resolved with supportive treatment. These data mirror those of the previously reported RAB/DOX study, and support the finding that alternating RAB/DOX is a reasonable treatment option for canine lymphoma.

Vincristine-induced adverse events related to body weight in dogs treated for lymphoma.

BACKGROUND: Traditional dosing of chemotherapy drugs based on body surface area may overdose small dogs, leading to an increased frequency of adverse events (AEs). HYPOTHESIS/OBJECTIVES: Evaluate the frequency of hematologic and gastrointestinal AEs in dogs with newly diagnosed lymphoma treated with vincristine weighing ≤15 kg in comparison to dogs weighing >15 kg. We hypothesized that dogs weighing ≤15 kg would experience a higher frequency of AEs. ANIMALS: One hundred and thirty-eight dogs with newly diagnosed lymphoma were treated with vincristine. METHODS: A multicenter retrospective study reviewing hematologic data and medical record information. Complete blood counts were performed no more than 24 hours before vincristine administration and then between 4 and 8 days post-administration. Data were evaluated using logistic regression or ordinal logistic regression. RESULTS: Thirty-eight dogs weighing ≤15 kg and 100 dogs weighing >15 kg were included. The median vincristine dose for both groups was 0.6 mg/m2. Seventeen (12.3%) instances of neutropenia occurred with no significant difference in overall frequency or grade between groups. Thirty initially asymptomatic substage A dogs (29.4%) experienced gastrointestinal AEs. Because of the widespread use of gastrointestinal supportive care medications, statistical comparison between groups could not be performed. Seven instances of hospitalization occurred (5.0%) and the risk of hospitalization did not differ significantly between groups (P = .37). CONCLUSIONS AND CLINICAL IMPORTANCE: Vincristine dosed at ≤0.6 mg/m2 does not increase the risk of hematologic AEs in dogs weighing ≤15 kg.

Jaundice caused by primary common bile duct lymphoma of a cat.

A 10-year-old American Shorthair cat presented with anorexia and jaundice, and echogenic evaluation revealed diffuse thickening of the common bile duct (CBD) wall. An exploratory laparotomy was conducted, the lesion was evaluated as difficult to remove, and the cat was euthanized and autopsied. Histologically, round neoplastic cells proliferated in the mucosa of the CBD and infiltrated the hepatic lobe, pancreas, and duodenum. Immunohistochemistry revealed that the neoplastic cells were positive for cytoplasmic-CD3 and granzyme B, and TCR-gamma clonal rearrangement was detected. Based on these findings, the neoplasia was diagnosed as a primary CBD lymphoma originating from cytotoxic T or natural killer cells. To the best of our knowledge, this is the first reported case of feline primary CBD lymphoma. Although rare, lymphoma of the CBD should be considered in cats with jaundice and thickening of the CBD.

Comparison of chemotherapy outcomes between normal and high serum cortisol concentration in dogs with lymphoma.

BACKGROUND: Increased serum cortisol (COR) concentrations may induce glucocorticoid resistance by down-regulation of glucocorticoid receptor (GCR), resulting in decreased chemotherapy efficacy in dogs with lymphoma. HYPOTHESIS: Investigate the relationship between serum COR concentrations and chemotherapy outcomes in dogs with lymphoma. ANIMALS: Thirty client-owned dogs with lymphoma, with serum COR concentration measured using serum samples collected at diagnosis. METHODS: Retrospective study. Dogs were divided into 2 groups based on serum COR concentrations: a normal group (n = 16) with COR concentrations <6 µg/dL and a high group (14) with COR concentrations ≥6 µg/dL. We compared signalment, clinical signs, stage, type of lymphoma, adrenal gland size, alkaline phosphatase (ALP) activity, response to chemotherapy, progression-free survival (PFS), overall survival (OS), and rate of P-glycoprotein (P-gp)- and GCR-positive cells between the 2 groups. RESULTS: No significant differences were found in the demographic characteristics between the 2 groups. However, the high COR group exhibited a significantly lower response to chemotherapy, PFS, and OS compared with the normal COR group. Serum ALP activity was significantly higher in the high COR group than in the normal COR group. Adrenal gland size was also significantly larger in the high COR group. Although no significant differences were found in the rate of P-gp-positive cells between the 2 groups, the rate of GCR-positive cells was significantly lower in the high COR group. CONCLUSIONS AND CLINICAL IMPORTANCE: Our data suggests that measurement of serum COR concentrations may serve as a potential prognostic factor and evaluation index.

Environmental radon, fracking wells, and lymphoma in dogs.

BACKGROUND: Multicentric lymphoma (ML) in dogs resembles non-Hodgkin lymphoma (NHL) in humans. Human NHL is associated with multiple environmental exposures, including to radon and volatile organic compounds (VOCs). HYPOTHESIS/OBJECTIVES: The objective of this study was to determine whether ML in dogs was associated with environmental radon or proximity to horizontal oil and drilling (fracking), a source of VOC pollution. METHODS: We identified dogs from the Golden Retriever Lifetime Study that developed ML (n = 52) along with matched controls (n = 104). Dog home addresses were categorized by Environmental Protection Agency radon zone and average residential radon by county, as well as by distance from fracking and associated wastewater wells. RESULTS: We found no significant differences in county level radon measurements. Individual household radon measurements were not available. There was no difference in residential proximity to active fracking wells between dogs with ML and unaffected dogs. While dogs with ML lived closer to wastewater wells (123 vs 206 km; P = .01), there was no difference in the percentage of cases vs controls that lived in close proximity (20 km) to a fracking well (11.5% for cases, 6.7% for controls; OR 1.81, 95% CI 0.55 to 5.22; P = .36), or a wastewater well (6.7% for cases, 4.4% for controls; P > .99). CONCLUSIONS AND CLINICAL IMPORTANCE: These data suggest that more proximate sources of chemical exposures need to be assessed in dogs with ML, including measurements of individual household radon and household VOC concentrations.

Spinal lymphoma in a goat.

A 3-year-old Pygmy Wether was presented for chronic hindlimb paralysis. A neurological exam revealed nonambulatory paraplegia with absent deep pain nociception, lack of hindlimb withdrawal reflexes, and paraspinal pain on palpation with T3 to L3 neurolocalization. MRI of the lumbar spine revealed an extensive, dorsal to dorsolateral, severely compressive, heterogeneously contrast-enhancing extradural lesion of the lumbar spine with intervertebral foraminal extension into the surrounding paraspinal musculature. Vertebral bone marrow involvement was also noted in the L5 and L6 vertebrae. A diagnosis of lymphoma was obtained after cytological sampling. This is the first case report describing specific MRI findings (signal characteristics, enhancement pattern, and perilesional changes) in a goat with paraspinal lymphoma.

Feline high-grade and large granular lymphocyte alimentary lymphomas treated with COP- or CHOP-based chemotherapy: A multi-centric retrospective study of 57 cases.

Specific data regarding outcome of cats with high-grade and large granular lymphocyte alimentary lymphoma (HGAL and LGL, respectively) treated with multi-agent chemotherapy are scarce. The aims of this multi-centric, retrospective study were to describe the outcome of cats with HGAL and LGL treated with COP- or CHOP-based chemotherapy and to identify potential prognostic factors. Cats with a cytological or histological diagnosis of HGAL or LGL lymphoma treated with COP- or CHOP-based protocol as first-line chemotherapy were included. Data regarding diagnosis, staging, treatment and follow-up were collected. Fifty-seven cats treated with CHOP (n = 37) or COP (n = 20) protocols were included. Complete response (CR), partial response (PR), stable disease (SD) and progressive disease (PD) were observed in 20%, 22%, 36% and 22% of cats, respectively, for an overall response rate of 42%. Median progression-free interval (PFI) was 148 days and overall median survival time (OST) was 131 days. Cats achieving CR, PR or SD showed significantly longer PFI (p < .01) and OST (p < .015) compared with cats with PD. Other positive prognostic factors in multi-variate analysis were rescue treatment (p < .001) and absence of lymph node involvement (p < .03). Negative prognostic factors were diffuse infiltration of the gastrointestinal tract (p = .035) and infiltration of a non-haematopoietic organ (p < .01).

Spindle cell cutaneous epitheliotropic T-cell lymphoma in an American Bulldog.

An 8-year-old American Bulldog developed coalescing exophytic bulbous nodules that grew rapidly on the left pinna and a single cutaneous mass on the left flank. Histological examination of the pinnal biopsy by a diagnostic laboratory revealed a densely cellular neoplasm with haphazardly arranged round to spindle cells with high mitotic activity and epitheliotropism. The initial diagnosis was a poorly differentiated malignant neoplasm with differential diagnoses including melanoma, tumour of histiocytic origin and, less likely, a pleomorphic lymphoma. A panel of melanoma immunohistochemical markers and immunolabelling for CD18 were pursued. Neoplastic cells were immunopositive for CD18 but negative for Melan-A, PNL2, TRP-1 and TRP-2, suggestive of a histiocytic tumour or lymphoma. The left ear masses recurred, and more masses developed on the body. The pinnectomized ear was submitted to the University of Missouri Veterinary Medical Diagnostic Laboratory. Similar cells were seen and were immunolabelled for CD18 and CD3 but were immunonegative for SOX10, CD79a and CD20. PCR for antigen receptor rearrangements revealed a clonal rearrangement of T-cell receptor gamma. These findings enabled a final diagnosis of epitheliotropic T-cell lymphoma with spindle cell morphology. Lymphoma should be considered as a potential differential diagnosis for cutaneous nodules of spindle cell morphology and lymphocytic immunohistochemical markers should be included in diagnostic panels.

In vitro chemosensitivity testing of the feline large granular lymphocyte cell line (S87).

BACKGROUND: Feline large granular lymphocyte (LGL) lymphoma is an aggressive neoplasia characterised by short survival and poor response to chemotherapy. OBJECTIVES: In this study, the effect of different chemotherapeutic agents on the growth kinetics of the feline cell line S87, a non-MHC-restricted feline LGL cell line, was investigated. Where possible, IC50 (inhibitory concentration 50) values were determined. The IC50 values of the cell line as lymphoma models can provide clues to the situation in vivo and serve as a basis for studying resistance mechanisms. METHODS: Cells were incubated with various concentrations of vincristine, doxorubicin, 4-hydroperoxycyclophosphamide, prednisolone, methotrexate and L-asparaginase for 24 and 48 h, respectively. RESULTS: The IC50 values could be determined as 14.57 (7.49-28.32) µg/mL at 24 h incubation and 5.72 (4.05-8.07) µg/mL at 48 h incubation for doxorubicin and 9.12 (7.72-10.76) µg/mL at 24 h incubation and 4.53 (3.74-5.47) µg/mL at 48 h incubation for 4-hydroperpoxycyclophosphamide. Treatment with vincristine and methotrexate resulted in relatively high cell resistance whereas L-asparaginase and prednisolone treatment led to a reduction in cell number compared to control while cell viability was not affected (cytostatic effect). CONCLUSION: Overall, the feline LGL cell line S87 proves to be relatively sensitive to doxorubicin and 4-hydroperoxycyclophosphamide and relatively resistant to treatment with vincristine, prednisolone, methotrexate and L-asparaginase. The results of this study can be used for further investigations on resistance mechanisms in feline LGL lymphoma. Doxorubicin and cyclophosphamide can be interpreted as promising candidates for the therapy of feline LGL lymphomas.

Assessment of Y chromosome copy number alterations in non-neoplastic and neoplastic leukocytes of male dogs.

The loss of the Y chromosome (ChrY), also known as LOY, is a common genetic alteration observed in men. It occurs in non-neoplastic cells as an age-related change as well as in neoplastic cells of various cancer types. While well-documented in humans, LOY has not been extensively studied in non-human mammals. In this study, we developed simple digital PCR-based assays to assess the copy number of ChrY relative to the X chromosome (ChrX) and chromosome 8 (Chr8) to evaluate ChrY numerical alterations in male canine DNA specimens. Using these assays, we analyzed non-neoplastic leukocytes from 162 male dogs without hematopoietic neoplasia to investigate the occurrence of age-related LOY in non-neoplastic leukocytes. Additionally, we examined 101 tumor DNA specimens obtained from male dogs diagnosed with various types of lymphoma and leukemia to determine whether copy number alterations of the ChrY occur in canine hematopoietic cancers. Analysis of the 162 non-neoplastic leukocyte DNA specimens from male dogs of varying ages revealed a consistent ∼1:1 ChrY:ChrX ratio. This suggests that age-related LOY in non-neoplastic leukocytes is rare or absent in dogs. Conversely, a decreased or increased ChrY:ChrX ratio was detected in canine neoplastic leukocytes at varying frequencies across different canine hematopoietic malignancies (P = 0.01, Fisher's exact test). Notably, a higher incidence of LOY was observed in more aggressive cancer types. To determine if this relative LOY to ChrX was caused by changes in ChrY or ChrX, we further analyzed their relative copy numbers using Chr8 as a reference. Loss of ChrX relative to Chr8 was found in 21% (9/41) of B-cell lymphomas and 6% (1/18) of non-T-zone/high-grade T-cell lymphomas. In contrast, a subset (29%, 4/14) of T-cell chronic lymphocytic leukemia showed gain of ChrX relative to Chr8. Notably, no relative LOY to Chr8 was detected indolent hematopoietic cancers such as T-zone lymphoma (0/9) and chronic lymphocytic leukemia of B-cell (0/11) and T-cell origins (0/14). However, relative LOY to Chr8 was present in more aggressive canine hematopoietic cancers, with incidences of 24% (10/41) in B-cell lymphoma, 44% (8/18) in non-T-zone/high-grade T-cell lymphoma, and 75% (6/8) in acute leukemia. This study highlights both similarities and differences in LOY between human and canine non-neoplastic and neoplastic leukocytes. It underscores the need for further research into the role of ChrY in canine health and disease, as well as the significance of LOY across various species.

Multicentric lymphoma in a donkey with intestinal and bone marrow involvement.

BACKGROUND: Lymphoma is a common neoplasm in horses but is reported much less commonly in donkeys. In this case report, we describe the macroscopic, microscopic and immunohistochemical features of a multicentric lymphoma with intestinal and bone marrow involvement. CASE PRESENTATION: A geriatric female donkey with history of chronic lameness was found dead. Post-mortem examination revealed advanced emaciation, periodontal disease, left front foot laminitis and multiple, soft, white to yellow tan intestinal transmural masses, up to 12 cm in diameter. Cytology suggested a round cell intestinal neoplasm. The femur of the left hint limb was double the size of the normal contralateral, with suspected neoplastic infiltration and replacement of bone marrow and bone. Histologically we diagnosed a lymphoma in the intestine and left femur. Immunohistochemically, the neoplastic cells showed CD3 immunolabelling, supporting a diagnosis of a multicentric T-cell lymphoma. CONCLUSIONS: To the authors' knowledge, this is the first time multicentric lymphoma is diagnosed in donkeys. Further studies of the genetic background, clinical, laboratory, histopathologic, and immunohistochemical, as well as the pathogenesis of lymphoma, is needed to better understand the uniquely low frequency of this neoplasm in donkeys.

Myeloid derived suppressor cells in peripheral blood can be a prognostic factor in canine transitional cell carcinoma.

Myeloid-derived suppressor cells (MDSCs) are immature cells with immunosuppressive properties found in the tumor microenvironment. MDSCs are divided into two major subsets: polymorphonuclear MDSCs (PMN-MDSCs) and monocytic MDSCs (M-MDSCs). Both MDSC subsets contribute to the creation of an immunosuppressive environment for tumor progression. In humans, patients with high levels of MDSCs show worse outcomes for several types of cancers. However, the association between MDSCs and clinical features has rarely been investigated in canine studies. In the present study, we measured the proportion of PMN-MDSCs and M-MDSCs in the peripheral blood and tumor tissue of dogs with hepatocellular carcinoma (HCC), prostate cancer (PC), transitional cell carcinoma (TCC), lymphoma, and pulmonary adenocarcinoma. Additionally, we examined immunosuppressive ability of PMN-MDSCs and M-MDSCs in peripheral blood mononuclear cells of TCC case on CD4+, CD8+ and interferon-γ+ cells and investigated the relationships of MDSCs with clinical features and outcomes. PMN-MDSCs increased in HCC, PC, TCC, and lymphoma. In contrast, M-MDSCs increased in the TCC. Both PMN-MDSCs and M-MDSCs exhibited immunosuppressive effects on CD8+, CD4+ and interferon-γ+ cells. In dogs with TCC, lymph node metastasis was associated with high level of PMN-MDSCs but not with M-MDSCs. High levels of both PMN-MDSCs and M-MDSCs were related to advanced tumor stage. Kaplan-Meier analysis revealed that high levels of both PMN-MDSCs and M-MDSCs were significantly associated with shorter overall survival. In addition, the Cox proportional hazard regression model showed that M-MDSCs and the tumor stage were independent prognostic factors for TCC. These results suggest that PMN-MDSCs and M-MDSCs may be involved in tumor progression and could be prognostic factors and promising therapeutic targets in dogs with TCC.

Plasma cell-free DNA in canine lymphoma patients as a novel material for genotyping.

Canine lymphoma is a disease with high morbidity and poor long-term prognosis, despite a high response rate to chemotherapy. In this study, we focused on liquid biopsy, in which small amounts of substances from body fluids were analysed, to determine whether cell-free DNA (cfDNA) in the plasma can be used as a biomarker for lymphoma in dogs. We found that 23 patients with lymphoma had significantly higher cfDNA concentrations than the 12 healthy dogs (median 2360 ng/mL versus 299 ng/mL, p < .0001). Polymerase chain reaction for antigen receptor rearrangement (PARR) was also employed using cfDNA from the lymphoma group to investigate whether cfDNA could be used for the detection of genetic clonality of lymphomas, as well as the genomic DNA (gDNA) extracted from an original lesion in each case. The correlation of the PARR results between cfDNA and gDNA was observed in 100% of B-cell lymphomas (10/10), 77.8% of T-cell lymphomas (7/9), and 100% of other types of lymphomas (4/4), respectively. These results indicate that plasma cfDNA levels are increasing in canine lymphoma patients, that cfDNA concentration can be a novel diagnostic tool, and that it can be used as a diagnostic tool for PARR.

Reply to Piewbang et al. Domestic Cat Hepadnavirus Antigens in Lymphoma Tissues. Comment on "Beatty et al. Domestic Cat Hepadnavirus and Lymphoma. Viruses 2023, 15, 2294".

We are grateful to the authors for providing additional data to demonstrate the presence of domestic cat hepadnavirus in lymphoma tissues [...].

Domestic Cat Hepadnavirus Antigens in Lymphoma Tissues. Comment on Beatty et al. Domestic Cat Hepadnavirus and Lymphoma. Viruses 2023, 15, 2294.

We are addressing the comments made by Beatty et al [...].