Background: Inflammatory mammary cancer (IMC), the counterpart of human inflammatory breast cancer (IBC), is the deadliest form of canine mammary tumors. IMC patients lack specific therapy and have poor outcomes. This proof-of-principle preclinical study evaluated the efficacy, safety, and effect on survival of neoadjuvant intratumoral (in situ) empty cowpea mosaic virus (eCPMV) immunotherapy in companion dogs diagnosed with IMC. Methods: Ten IMC-bearing dogs were enrolled in the study. Five dogs received medical therapy, and five received weekly neoadjuvant in situ eCPMV immunotherapy (0.2–0.4 mg per injection) and medical therapy after the second eCPMV injection. Efficacy was evaluated by reduction of tumor growth; safety by hematological and biochemistry changes in blood and plasma; and patient outcome by survival analysis. eCPMV-induced immune changes in blood cells were analyzed by flow cytometry; changes in the tumor microenvironment were evaluated by CD3 (T lymphocytes), CD20 (B lymphocytes), FoxP3 (Treg lymphocytes), myeloperoxidase (MPO; neutrophils), Ki-67 (proliferation index, PI; tumor cell proliferation), and Cleaved Caspase-3 (CC-3; apoptosis) immunohistochemistry. Results: Two neoadjuvant in situ eCPMV injections resulted in tumor shrinkage in all patients by day 14 without systemic adverse events. Although surgery for IMC is generally not an option, reduction in tumor size allowed surgery in two IMC patients. In peripheral blood, in situ eCPMV immunotherapy was associated with a significant decrease of Treg⁺/CD8⁺ ratio and changes in CD8⁺Granzyme B⁺ T cells, which behave as a lagging predictive biomarker. In the TME, higher neutrophilic infiltration and MPO expression, lower tumor Ki-67 PI, increase in CD3⁺ lymphocytes, decrease in FoxP3⁺/CD3⁺ ratio (p<0.04 for all comparisons), and no changes in CC-3⁺ immunostainings were observed in post-treatment tumor tissues when compared with pretreatment tumor samples. eCPMV-treated IMC patients had a statistically significant (p=0.033) improved overall survival than patients treated with medical therapy. Conclusions: Neoadjuvant in situ eCPMV immunotherapy demonstrated anti-tumor efficacy and improved survival in IMC patients without systemic adverse effects. eCPMV-induced changes in immune cells point to neutrophils as a driver of immune response. Neoadjuvant in situ eCPMV immunotherapy could be a groundbreaking immunotherapy for canine IMC and a potential future immunotherapy for human IBC patients.

The lack of optimal models to evaluate novel agents is delaying the development of effective immunotherapies against human breast cancer (BC). In this prospective open label study, we applied neoadjuvant intratumoral immunotherapy with empty cowpea mosaic virus-like particles (eCPMV) to 11 companion dogs diagnosed with canine mammary cancer (CMC), a spontaneous tumor resembling human BC. We found that two neoadjuvant intratumoral eCPMV injections resulted in tumor reduction in injected tumors in all patients and in noninjected tumors located in the ipsilateral and contralateral mammary chains of injected dogs. Tumor reduction was independent of clinical stage, tumor size, histopathologic grade, and tumor molecular subtype. RNA-seq-based analysis of injected tumors indicated a decrease in DNA replication activity and an increase in activated dendritic cell infiltration in the tumor microenvironment. Immunohistochemistry analysis demonstrated significant intratumoral increases in neutrophils, T and B lymphocytes, and plasma cells. eCPMV intratumoral immunotherapy demonstrated antitumor efficacy without any adverse effects. This novel immunotherapy has the potential for improving outcomes for human BC patients.

Canine inflammatory mammary cancer (IMC) is a highly aggressive and lethal cancer in dogs serving as a valuable animal model for its human counterpart, inflammatory breast cancer (IBC), both lacking effective therapies. Intratumoral immunotherapy (IT-IT) with empty cowpea mosaic virus (eCPMV) nanoparticles has shown promising results, demonstrating a reduction in tumor size, longer survival rates, and improved quality of life. This study compares the transcriptomic profiles of tumor samples from female dogs with IMC receiving eCPMV IT-IT and medical therapy (MT) versus MT alone. Transcriptomic analyses, gene expression profiles, signaling pathways, and cell type profiling of immune cell populations in samples from four eCPMV-treated dogs with IMC and four dogs with IMC treated with MT were evaluated using NanoString Technologies using a canine immune-oncology panel. Comparative analyses revealed 34 differentially expressed genes between treated and untreated samples. Five genes (CXCL8, S100A9, CCL20, IL6, and PTGS2) involved in neutrophil recruitment and activation were upregulated in the treated samples, linked to the IL17-signaling pathway. Cell type profiling showed a significant increase in neutrophil populations in the tumor microenvironment after eCPMV treatment. These findings highlight the role of neutrophils in the anti-tumor response mediated by eCPMV IT-IT and suggest eCPMV as a novel therapeutic approach for IBC/IMC.

Introduction: The clinical effect of low-level laser therapy (LLLT) on canine wounds is still under debate. The aim of this pilot study was to evaluate the potential influence of LLLT on the bacterial loads of wounds, using two different energy densities or doses of laser light as an adjuvant therapy for traumatic contaminated wound management. Methods: A prospective, randomized, blinded, placebo-controlled pilot clinical trial was used to evaluate the effect of two different doses of LLLT as an adjuvant treatment of contaminated traumatic wounds on the bacterial load and wound scoring in dogs. Fourteen dogs with traumatic bites or laceration wounds were randomly assigned to one of the three groups. Animals in groups A and B received a dose of LLLT of 6 and 2 J/cm2 respectively. Four wavelengths were used simultaneously: 660 nm, 800 nm, 905 nm, and 970 nm. Animals in group C received placebo LLLT. Bacterial burden and clinical wound scores were evaluated. Results: A statistically significant reduction in the average count of colony forming units was observed in group B (2 J/cm2 ) when compared to placebo group C. Group B also showed improved wound scores. No clinically adverse effects were observed in the patients treated with LLLT. Conclusion: LLLT, with the parameters used in this pilot trial, decreased bacterial loads of contaminated wounds in dogs and improved wound scores, especially when using a dose of 2 J/ cm2 . This is the first time the effect of LLLT on bacterial load has been investigated in a clinical setting using traumatic wounds in canine patients.

The incidence of surgical site infection (SSI) following mastectomy in female dogs remains poorly characterized, despite its relevance to postoperative outcomes and patient welfare. This retrospective cohort study aimed to determine the incidence of SSI and identify associated clinical and surgical risk factors in dogs undergoing mastectomy procedures at a veterinary teaching hospital between 2013 and 2022. SSI diagnosis was based on the Centers for Disease Control (CDC) criteria and identified structured passive surveillance of medical records. Univariate logistic regression was carried out to screen for potential risk factors, followed by multivariate analysis of selected variables. Among 376 mastectomy procedures performed in 306 dogs, 33 SSIs were detected, resulting in an overall incidence of 8.8% (95% CI: 6.1–12.1). The majority were superficial infections (93.9%), with only two cases classified as deep SSI; no organ/space infections were recorded. Prolonged anaesthesia duration (p = 0.041) and intraoperative hypothermia (p = 0.026) were independently associated with increased SSI risk. These findings indicate that the incidence of SSI after canine mastectomy is comparable to that reported for other clean surgeries. Prolonged anaesthesia and hypothermia represent modifiable risk factors, highlighting the importance of perioperative temperature control and optimized anaesthetic management. No significant associations were observed with patient-related variables, mastectomy type, or wound soaker catheter use, supporting its safe inclusion in multimodal analgesia.