Programmed Death-Ligand 1 (PD-L1) is an immune-checkpoint molecule involved in tumor-induced immunotolerance. While few studies explored its role in canine B-cell lymphoma (BCL), comprehensive analysis across lymphoma immunophenotypes remains limited. This study investigated associations between time to progression (TTP) and lymphoma-specific survival (LSS) with PD-L1 expression, including surface membrane protein (mPD-L1), mRNA levels in nodal aspirates, and soluble protein (sPD-L1) plasmatic concentrations, in dogs with different nodal lymphoma immunophenotypes at initial presentation. Fifty-eight cases were evaluated: 38 BCL, 11 T-cell lymphomas not otherwise specified (T-NOS), and 9 T-zone lymphomas (TZL). No significant association was found between mPD-L1 expression, mRNA level, sPD-L1 concentration, and TTP or LSS. Although not significant, median TTP was longer in mPD-L1 negative cases in both BCL (306, 95%C.I. 130-482 vs. 136, 95%C.I. 102-170 days) and T-NOS (228, 95%C.I. 40-416 vs. 48, 95%C.I. 0-168 days). The same was true for LSS (273, 95%C.I. 58-386 vs. 191, 95%C.I. 23-236 days for BCL, and 282, 95%C.I. 60-504 vs. 121, 95%C.I. 77-165 days for T-NOS). Notably, the only two TZL cases that progressed were mPD-L1-positive. These findings suggest a potential prognostic role for mPD-L1, warranting further validation in larger cohorts in BCL and T-NOS nodal lymphomas. For TZL, extended follow-up studies are needed due to its indolent behavior. Although sPD-L1 and mRNA levels did not correlate with outcome, their diagnostic and prognostic utility merits further investigation and methodologic refinement.

Elective neck dissection (END) and sentinel lymph node biopsy (SLNB) are suggested for nodal staging of canine head and neck malignancies (HNM). This study aims to compare the morbidity of END to SLNB. Seventy-six client-owned dogs with HNM that underwent END (n = 28) or SLNB (n = 48) in two institutions were retrospectively enrolled. Retrieved variables included data on signalment, lymph centre and lymph nodes, intra- and post-surgical complications (PSC) of lymphadenectomy, and histopathology results. The cumulative incidence of PSC at 30 days was estimated for END and SLNB and compared with Gray's test. The influence of variables on the incidence of complications was evaluated using univariate and multivariate models. No intraoperative complication occurred. The PSC were mostly mild. Seroma was the most frequent. The cumulative incidence of PSC of lymphadenectomy at 30 days was 47.4%, and they were severe in 14% of cases. The incidence of PSC was 25% for SLNB and 85.7% for END, and the difference was statistically significant (p < 0.001). Clinically enlarged nodes (p = 0.03), institution (p = 0.03), increasing number of resected nodes (p < 0.001) and of lymph centres (p < 0.001) predicted a higher incidence of PSC in the univariate model. In the multivariate analysis, only the type of node management (END vs. SLNB) remained significant. Although lymphadenectomy is a well-tolerated procedure in dogs with HNM, END was correlated with a higher risk of PSC compared to SLNB. Stratification of dogs by the risk of multiple nodal metastases is warranted to identify those who may still benefit from END despite a higher PSC risk.

Antibody-drug conjugates have transformed the treatment of cancer, yet their clinical utility can be limited by suboptimal tumor penetration, complex manufacturing, and safety concerns. Replacing antibodies with low-molecular-weight ligands enables the generation of small molecule-drug conjugates with enhanced tumor-targeting performance. Here, we report the results of a proof-of-concept study in canine cancer patients with OncoFAP glidotin, a small molecule-based targeted cytotoxic directed against Fibroblast Activation Protein. OncoFAP glidotin was evaluated in a dose-escalation trial in eight client-owned pet dogs bearing spontaneous FAP-positive tumors. The agent displayed an excellent safety profile, with no severe treatment-related adverse events. Six (75%) patients responded to the therapy. A tumor volume reduction of up to ∼88% in target lesions was observed on whole-body computed tomography, after only 4 weeks of treatment. These findings support further translational development activities of OncoFAP glidotin for both human and veterinary cancer patients.

The aim of this pilot study was to evaluate the feasibility and utility of real-time tumor fluorescence-guided surgery (TFGS) with near-infrared fluorescence indocyanine green (NIRF-ICG) for the removal of spontaneous soft tissue sarcoma (STS) and mast cell tumor (MCT) at first presentation or recurrence in dogs. A dose of 0.5 mg/kg of ICG was administered 24 h before surgery, with a tumor-to-background (TBR) and signal-to-background ratio (SBR) cut-off of 2. Twenty-four dogs with 28 tumors (14 MCTs and 14 STSs) were included. TFGS was feasible in all STSs and in 11/14 MCTs. The median TBR of STS and MCT was 13 (IQR 26–9.4) and 5.8 (IQR 19–3.6) respectively. Considering the residual fluorescence in the wound bed and the histopathological results the sensitivity and specificity of NIRF-ICG excision were 80% and 78% in STS and 60% and 50% in MCT, respectively. Overall, resection was extended during TFGS in 64% of STSs and 55% of MCTs. These results convey the feasibility of TFGS with NIRF-ICG in canine STS surgical extirpation. However, the findings indicate more limited performance in MCT. The different performance in STS and MCT needs to be investigated in a larger population.

No data are available about the sentinel lymph node (SLN) mapping in presence of enlarged regional lymph nodes (eRLN), due to metastatic or inflammatory processes. The present study aims to assess the influence of eRLN clinically suspicious for nodal metastasis in SLN mapping in canine malignancies and describe possible alterations of the nodal lymphatic drainage and tracer uptake.Dogs with malignancies and eRLN were included and underwent to SLN mapping with lymphoscintigraphy and/or near-infrared fluorescence (NIRF). Findings in SLN mapping, distribution of nodal tracer uptake and histological nodal status were recorded.Twenty-two dogs with eRLN were included. During the lymphographies 2 patterns of nodal tracer distribution were observed: in 9 lymphographies (41%) only the eRLN (single or multiple) was identified (Pattern 1); in 13 (59%), beside the eRLN, at least one non-palpable/normal-sized SLN was identified (Pattern 2). The adjunctive SLNs were detected in the same (25%) or in a different (75%) lymphocentrum of the eRLN. The 54% of the adjunctive non-palpable/normal-sized SLN were metastatic. Among the pattern 2, in 3 lymphographies the eRLN had incomplete or absent nodal distribution of tracer uptake and the tracers were rerouted to one or more non-palpable/normal-sized SLN becoming the neo-SLN.Neoplastic or inflammatory status of the lymph node may alter the lymphatic drainage in example leading to find non-palpable/normal-sized SLN and various distribution of tracers’ uptakes were recorded in eRLN. In presence of eRLN the SLN mapping is strongly suggested rather than limiting lymphadenectomy of only enlarged node, to avoid missing potentially residual microscopic neoplastic nodal disease in additional SLN.

IntroductionFibroblast activation protein (FAP) is involved in the extracellular matrix (ECM) remodeling and wound healing. Absent in most adult tissues, it is overexpressed by neoplastic cells and/or cancer-associated fibroblasts (CAFs) in several human malignancies. The extra Domain-B of fibronectin (EDB+FN) is a splice variant of fibronectin involved in angiogenesis and tissue remodeling, overexpressed by CAFs and cancer-associated vessels (CAVs) in many aggressive human tumors. This study aims to investigate FAP and EDB+FN expressions in canine tumors and assess their potential as druggable targets in animal patients.MethodsFAP and EDB+FN expression was assessed by immunohistochemistry on 88 canine tumors, including Soft Tissue Sarcomas [STS], Osteosarcomas [OSA], Hemangiosarcomas [HSA], Apocrine Gland Anal Sac Adenocarcinomas [AGASAC], Mast Cell Tumors [MCT], Lymphomas, and Melanomas, using polyclonal and monoclonal anti-FAP and the L19 anti-EDB antibodies. Expression distribution and intensity were semi-quantitatively scored in neoplastic cells, CAFs, CAVs, and stroma.ResultsFAP was variably expressed in neoplastic cells (79/88), CAFs (79/88), and CAVs (82/88) across all tumor types, but mostly in AGASACs, STSs, and MCTs. The monoclonal antibody presented greater specificity. EDB+FN expression was less present across tumor types, mostly with a vascular staining pattern. Labelling was most intense and consistent in the neoplastic cells, CAFs, and CAVs of melanomas, and to a lesser extent in AGASAC and STS.DiscussionSTS, AGASAC, and MCT could be candidates for FAP-targeted strategies; melanomas are the most promising for EDB+FN-directed therapies. These results support FAP and EDB+FN as targets worth investigating for clinical applications in animal patients.

Sentinel lymph node mapping is increasingly used in canine and feline oncology and often involves the combined use of visible dyes and fluorescent tracers. However, the effect of methylene blue on the fluorescence of indocyanine green during near-infrared imaging remains unclear. This explorative study aimed to quantitatively and qualitatively assess potential fluorescence quenching in solutions of methylene blue-indocyanine green at different ratios in three near-infrared imaging modalities (overlay, color map, contrast). Four solutions were prepared: 100%/0%, 75%/25%, 50%/50%, and 25%/75% indocyanine green/methylene blue. The fluorescence intensity of the four solutions was quantitatively measured in vitro using near-infrared imaging. Subsequently, four lymphographies, one for each solution, were performed from the metatarsal region of feline cadavers. Observers with varying levels of experience evaluated lymphographic images. Methylene blue caused a concentration-dependent reduction in fluorescence both at the quantitative evaluation and qualitative lymphography interpretation. Despite this reduction, fluorescence remained sufficient in cadavers for accurate identification of lymph nodes, and observer experience did not significantly affect interpretation, except for the color map mode. Because methylene blue-dominant solutions showed a greater quenching effect on indocyanine green fluorescence, clinicians should favor indocyanine green-dominant mixtures. This approach may preserve fluorescence performance, maintaining the surgical guidance benefits of methylene blue. Future confirmatory studies should include a substantially larger number of specimens to allow appropriate statistical comparisons and to better account for inter-individual variability.

IntroductionFlow cytometry (FC) has been used recently to assess percentages of infiltration by mast cells in sentinel lymph nodes (SLN) of dogs with mast cell tumors (MCT). SLN mapping often includes the use of different dyes such as methylene blue (MB) and indocyanine green (ICG), which might influence fluorescence assessment by FC. This study aimed to assess whether the color given by mapping dyes might affect the baseline fluorescence of SLN aspirates for FC.MethodsBaseline fluorescence was calculated as Median Fluorescence Intensity (MFI) in 4 channels (FL1, FL2, FL3, FL4, respectively corresponding to 530/30 nm, 585/40 nm, >670 nm, 660/20 nm) with a cytometer equipped with two lasers (488 nm and 638 nm) with constant setting and compensation. SLN aspirates were suspended in RPMI medium. They were classified based on results of mapping techniques in 4 dye classes (blue/fluorescent, fluorescent/non-blue, blue/non-fluorescent, non-blue/non-fluorescent), and possible differences in MFI values among SLN dye classes were assessed.ResultsThirty-five SLNs from 17 dogs were assessed. Considering dye classes, 13 were blue/fluorescent, 16 were fluorescent/non-blue, 4 were blue/non-fluorescent, 2 were non-blue/non-fluorescent. In all fluorescence channels, except FL1, MFI varied among SLN dye classes. Blue/non-fluorescent SLNs showed the highest fluorescence, followed by blue/fluorescent, fluorescent/non-blue, and non-blue/non-fluorescent.DiscussionThese results suggest that the use of dyes for SLN mapping may introduce a relevant bias when MFI is quantitatively assessed via FC.

Canine apocrine gland anal sac adenocarcinoma (AGASAC) is an aggressive malignancy with a high incidence of regional lymph node metastasis at diagnosis. Platelet-derived growth factor receptor beta (PDGFRβ) is a receptor tyrosine kinase involved in oncogenic signaling and angiogenesis, representing a potential therapeutic target. Its expression in different AGASAC histotypes has not been fully defined. This study evaluated microscopic patterns and PDGFRβ immunohistochemical expression in three normal anal gland sacs, 51 primary AGASACs (12 non-metastatic, 39 metastatic), and 33 corresponding nodal metastases. PDGFRβ expression was semi-quantitatively scored and statistically analyzed. Histotypes included 26 mixed with solid prevalence, 11 pure solid (including 1 comedo-carcinoma), 9 mixed with tubular prevalence, 4 tubular, and 1 neuroendocrine-like. PDGFRβ was expressed in normal anal sac glands and in 43/51 tumors (5–100% positive cells): 19/26 mixed with solid prevalence, 10/11 solid, 9/9 mixed with tubular prevalence, 4/4 tubular, and 1 neuroendocrine-like. PDGFRβ labeled 27/33 nodal metastases. PDGFRβ expression was highest and more intense in tumors with a tubular pattern. The statistical trend indicated a correlation of AGASAC dedifferentiation to reduced PDGFRβ expression, but no statistical significance was found. Rare AGASAC variants (solid comedo-carcinoma and neuroendocrine-like) were described for the first time. PDGFRβ expression was higher in tubular tumors and commonly detected in primary and metastatic lesions. These findings support exploring TKI therapy for specific AGASAC histotypes. Further studies integrating receptor activation and treatment outcomes are warranted to clarify predictive and prognostic relevance.

Serum activity of paraoxonase-1 (PON-1) decreases in canine inflammation. This study aimed to evaluate how frequently PON-1 activity is reduced across different disease categories and to assess its potential prognostic significance. PON-1 activity was measured in 482 samples (435 first visit, 47 follow-up) collected during routine clinical activities. The emergency/first-opinion unit (EF) had the highest frequency of low values (18.3%) and lower median PON-1 activity (170.3 U/mL) compared with other units. The proportion of lower values and median PON-1 values were, respectively, significantly higher and lower in acute/severe diseases compared with chronic/mild diseases (13.3 vs. 7.1%; 177.0 vs. 192.9 U/mL) and in hospitalized compared with non-hospitalized dogs (27.7 vs. 4.5%; 150.0 vs. 193.0 U/mL). Low PON-1 activity may predict the need for hospitalization: values < 45.7 U/mL have a likelihood ratio of 14.4. The proportion of lower values and the median Paraoxonase-1 activity did not differ between survivors and non-survivors (18.2 vs. 25.0%; 162.0 vs. 161.3 U/mL). However, PON-1 activity diminished during hospitalization only in non-survivors. PON-1 activity should be measured in routine practice, especially in the EF; hospitalization may be warranted when results are markedly low. Decreases in PON-1 activity during hospitalization may suggest a negative outcome.

To date, animal models for lymphographic studies mainly focused on dog, while lymphography is rarely reported in cats, and even less involving cutaneous lymphatic territories. This study aims to assess the feasibility of cutaneous lymphography using indocyanine green (ICG) fluorescence in cat cadavers and describe predictable lymphatic pathways from cutaneous regions of head and hind limb anatomical districts. Frozen or refrigerated cadavers of adult cats that died for causes unrelated to the study were included. Twenty cutaneous regions (6 from the head; 14 from the hind limb) were selected using easily assessable anatomical landmarks, and expected draining lymphocentrums were presumed based on canine studies since there is no similar information for cats. For each lymphography, a single selected cutaneous region per anatomical district was assessed. After intradermal ICG injections, lymphatic drainage was favored by massage and/or flexion-extension movements. For each lymphography, all expected and detected lymphocentrums were dissected, and lymph nodes extirpated. Variables regarding cadavers and lymphography characteristics were assessed. ICG-lymphography was repeated in 33 cadavers. Out of the 99 selected cutaneous regions available, 15 were excluded following inclusion criteria, therefore lymphographies were performed for a total of 84 selected cutaneous regions (26 from the head and 58 from the hind limbs). A success was recorded in 63/84 (75%) lymphographies, with a median migration time of 8 (1-30) minutes. The ICG drained to the expected lymphocentrum in 28/63 (44%) lymphographies, and to other ones in 35/84 (56%). ICG-lymphography is feasible in cat cadavers, regardless of technique or cadaver characteristics. The observed difference in lymphatic drainage (56% to unexpected lymphocentrums) highlights the importance of specifically mapping lymphatic territories in cats. ICG-lymphography demonstrated as an effective technique and could be used to improve knowledge of feline lymphatic physiology. Further studies may provide a more complete understanding of superficial lymphatic territories in cats.

BACKGROUND:No data regarding near-infrared fluorescence (NIRF) with indocyanine green and its comparison with lymphoscintigraphy are available for sentinel lymph node identification in canine and feline solid malignant tumors. This study compares both techniques in sentinel lymphocentrums mapping and surgical guided sentinel lymph node extirpation. RESULTS:Fifty-four animals with 60 tumors were included, and the combined use of lymphoscintigraphy and NIRF was applied: 80 sentinel lymphocentrums were surgically explored, and 113 sentinel lymph nodes were extirpated. This combination allowed the detection of at least one sentinel lymphocentrum and sentinel lymph node per each tumor. During mapping, the sentinel lymphocentrum detection rate with NIRF was 93%, similar to lymphoscintigraphy with a handheld intraoperative gamma probe (92%). No significant differences among techniques were observed in the surgical guided exploration phase, except that the number of fluorescent sentinel lymph nodes was significantly higher than radioactive ones, and the number of detected metastatic lymph nodes was comparable among the techniques. Surgeons subjectively considered the intraoperative gamma probe more helpful in 46.5% of sentinel lymph node extirpations, NIRF more helpful in 24.2%, and both techniques equally helpful in 29.3% of cases. Overall, they deemed the use of intraoperative gamma probe superior to NIRF for axillary lymphadenectomies and NIRF superior to lymphoscintigraphy for head and neck lymphadenectomies. CONCLUSIONS:Despite some differences between the techniques, NIRF with indocyanine green can be considered a practical and viable alternative to lymphoscintigraphy for sentinel lymphocentrum mapping and guided sentinel lymph node removal in small animal oncologic practice, particularly where lymphoscintigraphy is not accessible.

IntroductionCanine soft tissue sarcomas (STSs) are locally aggressive mesenchymal tumors with variable recurrence rates, and often, their therapy is limited to surgical excision. CD117 (KIT) is a tyrosine kinase receptor involved in cell growth and cancer development. c-kit proto-oncogene mutations have been reported to be associated with prognosis and therapy response in human and canine cancers. However, CD117 expression and c-kit mutations have rarely been investigated in canine STSs. This study aims to assess CD117 expression and c-kit mutations in different canine STSs.MethodsSpontaneous STSs were surgically removed, fixed, routinely processed, and stained for histological and anti-CD117 immunohistochemical analyses. Staining intensity and percentage of positivity were scored. Cases with intense CD117 expression in more than 50% of cells were analyzed for the presence of mutations in exons 8, 9, or 11 of the c-kit proto-oncogene.ResultsOverall, 115 canine STSs were collected. Among them, CD117 was expressed in 43 STSs, with diffuse cytoplasmic staining of variable intensity. CD117 was expressed in 16 out of 27 perivascular wall tumors, 12 of 13 sarcomas of fibroblastic origin, 6 of 6 rhabdomyosarcomas, 7 of 46 liposarcomas, and 2 of 3 nerve sheath tumors. Leiomyosarcomas (20 of 20) did not show CD117 expression. Mutations were investigated in 22 cases, all of which returned negative results.DiscussionIn summary, canine STSs variably expressed CD117, which suggests that tyrosine kinase inhibitors may represent a promising targeted therapy for selected canine STSs histotypes.

Abstract Background The therapeutic role and prognostic relevance of lymphadenectomy in mast cell tumor (MCT) has historically been evaluated on regional rather than sentinel lymph nodes. Hypothesis/Objectives To update information about the association of histological nodal (HN) classes with clinical outcome in dogs with MCT after tumor excision and extirpation of normal‐sized sentinel nodes (SLN) guided by radiopharmaceutical. Animals Ninety‐four dogs with histologically‐confirmed treatment‐naïve MCT (71 cutaneous, 22 subcutaneous and 1 conjunctival MCT) were included if without: distant metastases, lymphadenomegaly, concurrent mixed cutaneous, and subcutaneous MCT. Methods This was a monoistitutional cohort study. Tumors characteristics were retrieved and SLNs were classified according to Weishaar's system. Incidence of MCT‐related events (local, nodal, distant relapse), de novo MCT or other tumors and death (MCT‐related and non‐MCT‐related), were recorded. Incidence curves were compared among the HN classes. Results Twenty‐seven dogs had HN0, 19 HN1, 37 HN2, and 11 HN3 SLN. Thirteen (2 HN0, 4 HN2, and 7 HN3) received adjuvant chemotherapies. Kiupel high grade, increasing number of SLN and lymphocentrums were associated with higher HN classes. Five dogs died for MCT‐related causes: 1 low‐grade (HN0) and 1 subcutaneous (HN3) had a local relapse, 2 high‐grade had distant relapse (HN3‐HN0) and 1 dog developed disease progression from a de novo subcutaneous MCT. No nodal relapse was registered. Fourteen dogs developed de novo MCTs. Conclusion/Discussion Low grade/low‐risk MCT with nonpalpable and normal sized SLN have a favorable outcome independently from the HN. Result should be considered strictly related to the successful SLN detection guided pre‐ and intraoperative by radiopharmaceutical markers.

Due to the low frequency and the changes in diagnostic techniques and terminology during the last few years, only little clinical information is available on splenic stromal sarcoma (SSS). This multi-institutional study aimed at gathering clinical cases of SSS in dogs and investigates their clinical behaviour, as well as analyse possible clinicopathological prognostic factors, including the use of adjuvant therapy. Dogs with a histologically confirmed SSS that underwent splenectomy were retrospectively included. To be included in the study, either FFPE tissue blocks or multiple tissue sections had to be available for histopathologic and immunohistochemical revision. Clinical and pathological variables, along with adjuvant therapy data, were collected. Cumulative incidence of metastatic disease was analysed through univariate and bivariate analyses. The impact of adjuvant chemotherapy on metastasis incidence and survival was assessed, considering an estimated propensity score. A total of 32 dogs were included. Among them, 22 developed metastases with an incidence of 37.5%, 59.38%, and 65.94% at 6, 12, and 24 months, respectively. Univariate analysis identified mitotic count, total scoring, and necrosis as prognostic factors. In bivariate analysis, mitotic count remained prognostic. The administration of adjuvant chemotherapy did not have an impact on metastasis incidence or survival time. The study found that dogs with SSSs are at high risk of metastasis, although a small subgroup may experience longer survival after splenectomy. Mitotic count was the only variable having a reliable prognostic impact. Adjuvant chemotherapy did not appear to decrease the incidence of metastasis or prolong survival in these dogs.

(1) Background: the erythrocyte sedimentation rate (ESR) has been reported to increase in some infectious or inflammatory diseases in dogs, but no information on the frequency of increases in a routine clinical setting exists. The aim of this study was to assess the frequency of an increased ESR in dogs and to investigate its possible association with hematologic changes; (2) Methods: A total of 295 EDTA blood samples were randomly selected from the routine caseload of the Veterinary Teaching Hospital. Samples were grouped in controls and in pathologic groups based on the clinical presentation. A routine hemogram was performed, then the ESR was measured using the instrument MINI-PET; (3) Results: compared with controls, the ESR was significantly higher in all the pathologic groups, except for the hematological disorders group. The highest ESR was found in samples from dogs with chronic kidney disease or inflammation, followed by those from dogs with mild chronic disorders, severe/acute diseases, tumors and urinary disorders. The ESR negatively correlated with hematocrit and positively with neutrophil counts. (4) Conclusions: The ESR increases more frequently in dogs with clinically evident inflammation or CKD, but also in several other conditions, likely as a consequence of anemia and acute phase response.

Osteosarcoma is the most common malignant primary bone cancer, but it is infrequently reported in cats. Feline appendicular osteosarcoma typically exhibits good prognosis when treated with surgery alone. A retrospective multi-institutional study was conducted to identify possible prognostic factors. Cats diagnosed with appendicular osteosarcoma were included if initial staging and follow-up information were available. Data including signalment, tumour characteristics, treatment modalities, and survival outcomes were collected and analysed. Fifty-six cats were included; the femur was the most frequently affected bone. Eight cats had distant metastasis at admission and an additional 9 developed metastatic disease during follow-up, resulting in an overall metastatic rate of 30%. Forty-nine (87.5%) cats underwent surgery, and 4 also received adjuvant chemotherapy. Among operated cats, median time to local progression (TTLP), time to distant progression and tumour-specific survival (TSS) were not reached. One- and 2-year survival rates were 66% and 55%, respectively. Seven (12.5%) cats received no treatment; 1- and 2-year survival rates were 25% and 0%, respectively. Operated cats had significantly longer TTLP (P < .001) and TSS (P = .001) compared with non-operated cats. Among operated cats, young age negatively impacted local tumour progression, while the presence of distant metastasis at diagnosis was associated with a higher risk of tumour-related death. This study reaffirms the good prognosis for cats with appendicular osteosarcoma undergoing surgery, but sheds light on some additional factors to consider. Accurate initial staging is recommended, as the metastatic rate may exceed many previous estimations. Surgery substantially extends survival time, whereas the role of chemotherapy remains uncertain.

Cutaneous and subcutaneous mast cell tumors (MCTs) are common canine neoplasms characterized by variable biological behavior. Tumor-associated macrophages (TAMs) and tumor-infiltrating lymphocytes (TILs) can be effective prognostic markers in numerous human neoplasms and are increasingly investigated in dogs. The aim of this study was to characterize immune cells in canine MCTs and their relationship with histological location (cutaneous, subcutaneous) and histologic nodal metastatic status (HN0-3). Thirty-eight MCTs (26 cutaneous, 12 subcutaneous) from 33 dogs with known sentinel lymph node (SLN) metastatic status were immunolabeled for Iba1 (macrophages), CD20 (B cells), CD3 (T cells), and Foxp3 (regulatory T cells). Semiquantitative scoring of interstitial and perivascular CD3+, CD20+, and Foxp3+ cells and morphological evaluation of Iba1+ cells were performed. For each marker, the percent immunopositive area was evaluated by image analysis. All MCTs were diffusely infiltrated by Iba1+ cells and variably infiltrated by CD20+, CD3+, and rare Foxp3+ cells. Stellate/spindle Iba1+ cells were associated with HN2 and HN3 SLNs. Perivascular Foxp3+ cells, CD3+ cells, and percent CD3+ areas were increased in subcutaneous MCTs. Interstitial CD3+ cells were increased in cutaneous MCTs with HN0 SLNs. No differences in CD20+ cells were identified between cutaneous and subcutaneous MCTs and among SLN classes. MCTs were markedly infiltrated by TAMs and variably infiltrated by TILs. Stellate/spindle morphology of TAMs associated with HN2 and HN3 SLNs is suggestive of a pro-tumoral (M2) phenotype. Cutaneous and subcutaneous MCTs have different tumor-immune microenvironments, and T-cell infiltration might contribute to prevention of nodal metastatic spread of cutaneous MCTs.

High survivin expression has been correlated with poor outcomes in several canine tumors but not in soft tissue tumors (STTs). Survivin is a target gene of the Wnt/β-catenin pathway, which is involved in human STT oncogenesis. Immunohistochemistry for survivin, β-catenin, and Ki-67 was performed on 41 canine perivascular wall tumors (cPWTs), and statistical associations of protein expression and histopathologic and clinical variables with clinical outcomes were investigated. Immunohistochemically, there was nuclear positivity (0.9%–12.2% of tumor cells) for survivin in 41/41 (100%), cytoplasmic positivity (0 to > 75% of tumor cells) for survivin in 31/41 (76%), nuclear positivity (2.9%–67.2% of tumor cells) for β-catenin in 24/41 (59%), and cytoplasmic positivity (0% to > 75% of tumor cells) for β-catenin in 23/41 (56%) of cPWTs. All tumors expressed nuclear Ki-67 (2.2%–23.5%). In univariate analysis and multivariate analysis (UA and MA, respectively), every 1% increase of nuclear survivin was associated with an increase of the instantaneous death risk by a factor of 1.15 [hazard ratio (HR) = 1.15; P = .007]. Higher nuclear survivin was associated with grade II/III neoplasms ( P = .043). Expression of cytoplasmic survivin, nuclear and cytoplasmic β-catenin, and nuclear Ki-67 were not significantly associated with prognosis in UA nor MA. Tumor size was a significant prognostic factor for local recurrence in UA [subdistribution HR (SDHR) = 1.19; P = .02] and for reduced overall survival time in MA. According to UA and MA, a unitary increase of mitotic count was associated with an increase of the instantaneous death risk by a factor of 1.05 (HR = 1.05; P = .014). Nuclear survivin, mitotic count, and tumor size seem to be potential prognostic factors for cPWTs. In addition, survivin and β-catenin may represent promising therapeutic targets for cPWTs.