Ki67 Index in Patnaik Grade 2/Kiupel Low-Grade Canine Cutaneous Mast Cell Tumors with Early Lymph Node Metastasis: A Descriptive Study.

Several studies evaluating Ki67 in canine cutaneous mast cell tumors (cMCTs) have reported its prognostic value when tumors of all histological grades are included. This study aims to evaluate whether the Ki67 index has a predictive value in a homogeneous cohort of G2/LG cMCTs with HN2 lymph nodes (LNs) and to describe the clinical outcome. The second goal was to explore the correlation between the Ki67 index and MC. The medical databases of three institutions were retrospectively searched for dogs undergoing surgical treatment for cMCT and LN extirpation, with a histological diagnosis of G2/LG with HN2 LNs. Information about histological margins, MC, Ki67 index, local recurrence, nodal relapse, distant metastasis, de novo cMCT occurrence and date and cause of death were included. A total of 39 cases were identified. None of these developed local and nodal relapse or metastatic distant disease. Median MC was 1 (0-2). Median Ki67 index was 3.5 (0.7-14.3). Ki67 and MC were not significantly correlated. At the end of the study, 32 (82%) dogs were alive, 7 (18%) dogs were dead from unrelated causes and 4 (10.2%) dogs were lost to follow-up. The median ST was not reached, and the mean was 893 days (104-2241 days). Considering the strict inclusion criteria, dogs affected by G2/LG with HN2 LNs treated with surgery alone may have a good oncologic outcome; the Ki67 index does not have prognostic impact.

Development of Monoclonal Antibodies Targeting Canine PD-L1 and PD-1 and Their Clinical Relevance in Canine Apocrine Gland Anal Sac Adenocarcinoma.

Canine apocrine gland anal sac adenocarcinoma (AGASACA) is an aggressive canine tumor originating from the anal sac glands. Surgical resection, with or without adjuvant chemotherapy, represents the standard of care for this tumor, but the outcome is generally poor, particularly for tumors diagnosed at an advanced stage. For this reason, novel treatment options are warranted, and a few recent reports have suggested the activation of the immune checkpoint axis in canine AGASACA. In our study, we developed canine-specific monoclonal antibodies targeting PD-1 and PD-L1. A total of 41 AGASACAs with complete clinical and follow-up information were then analyzed by immunohistochemistry for the expression of the two checkpoint molecules (PD-L1 and PD-1) and the presence of tumor-infiltrating lymphocytes (CD3 and CD20), which were evaluated within the tumor bulk (intratumor) and in the surrounding stroma (peritumor). Seventeen AGASACAs (42%) expressed PD-L1 in a range between 5% and 95%. The intratumor lymphocytes were predominantly CD3+ T-cells and were positively correlated with the number of PD-1+ intratumor lymphocytes (ρ = 0.36; p = 0.02). The peritumor lymphocytes were a mixture of CD3+ and CD20+ cells with variable PD-1 expression (range 0-50%). PD-L1 expression negatively affected survival only in the subgroup of dogs treated with surgery alone (n = 14; 576 vs. 235 days). The presence of a heterogeneous lymphocytic infiltrate and the expression of PD-1 and PD-L1 molecules support the relevance of the immune microenvironment in canine AGASACAs and the potential value of immune checkpoints as promising therapeutic targets.

Use of a customized 3D-printed external splint for the correction of a severe pectus excavatum in a 3-month-old kitten.

Background: Pectus excavatum is a rare congenital or developmental deformity of the chest wall that has been reported in both dogs and cats. The clinical symptoms vary in severity and are correlated with the grade of the deformity. The most severe cases are at a very high risk of intra- and post-operative complications that could be fatal. Case Description: A 3-month-old kitten was presented for severe dyspnea and exercise intolerance. After the radiographic diagnosis of severe pectus excavatum, a splint was designed based on the computed tomography scan of the kitten. A locking mechanism was applied on a dedicated pocket on the splint and secured with commercial glue. Five sutures were placed, the most caudal three with a surgical approach to the corresponding sternebra. The three caudal traction sutures were progressively tensioned during the first 2 weeks, and then tied. The splint was removed 1 month postoperatively; neither surgical nor splint-associated complications were observed. The kitten well tolerated the splint and the owner reported no further episodes of dyspnea or exercise intolerance. Vertebral and frontosagittal indexes were 3.2 and 9.1 at presentation, respectively, and 10.2 and 2.0 at 4 months postoperatively. Conclusion: The use of a patient-specific 3D-printed external splint associated with a progressive correction of the pectus excavatum can be a better solution for the treatment of severe cases of pectus excavatum, where a sudden distension of the rib cage could cause re-expansion injuries.

Prognostic impact of bone invasion in canine oral malignant melanoma treated by surgery and anti-CSPG4 vaccination: A retrospective study on 68 cases (2010-2020).

Prognosis of canine oral malignant melanoma encompasses clinical, histological and immunohistochemical parameters. The aim of this study was to evaluate the prognostic impact of bone invasion in oral canine melanoma. Sixty-eight dogs bearing oral melanoma staged II and III that underwent surgery and anti-CSPG4 electrovaccination, with available histological data and a minimum follow up of minimum 1 year, were retrospectively selected. Bone invasion was detected on imaging and/or histology. Median survival time of dogs with evidence of bone invasion (group 1) was 397 days and significantly shorter compared with dogs with oral melanomas not invading the bone (group 2, 1063 days). Dogs with tumours localised at the level of the cheek, lip, tongue and soft palate (soft tissue - group 3) lived significantly longer compared with dogs having tumours within the gingiva of the maxilla or mandible (hard tissue - group 4) with a median survival time of 1063 and 470 days, respectively. Within group 4, the subgroup of dogs with tumours not invading the bone (group 5) showed a significant prolonged survival time (972 days) in comparison with dogs of group 1 (bone invasion group). Similar results were obtained for the disease-free intervals amongst the different groups. Statistical analysis showed that Ki67 and mitotic count were correlated with shorter survival in patients of group 1 (with bone invasion). Bone invasion should always be assessed since it appears to be a negative prognostic factor.

Canine Epithelial Thymic Tumors: Outcome in 28 Dogs Treated by Surgery.

Thymoma is a tumor rarely reported in dogs and should be differentiated from mediastinal lymphoma. Clinical signs may have a late onset, and thymoma is often diagnosed when symptoms related to the space-occupying effect or paraneoplastic syndromes occur. CT and fine-needle aspirates or core biopsies are helpful in differential diagnosis, but flow cytometry may improve the pre-operative diagnostic ability. Concurrent paraneoplastic syndromes such as myasthenia gravis and hypercalcemia have been reported; however, their role as prognostic factors is not well determined. Surgical excision is the treatment of choice; adjuvant radiotherapy and/or chemotherapy may prolong survival in cases of incomplete excision or when a thymic carcinoma is diagnosed. Local recurrence and metastasis are infrequently reported; therefore, a long survival time is expected if the tumor is completely excised or if adjuvant therapy is undertaken. This article reports the authors' experience with 28 dogs affected by 18 thymomas and 10 thymic carcinomas. The median overall survival in this series was 1173 days, and the median disease-free interval was 903 days. Dogs with thymic carcinoma had significantly shorter disease-free intervals and shorter, although not statistically significant, survival times. Dogs with Masaoka Stage III tumors had worse outcomes.

Prognostic Value of Ki67 and Other Clinical and Histopathological Factors in Canine Apocrine Gland Anal Sac Adenocarcinoma.

Apocrine gland anal sac adenocarcinoma (AGASACA) is locally aggressive and highly metastatic to regional lymph nodes. The aim of this study was to evaluate the prognostic significance of Ki67 in surgically excised AGASACA. Prognostic impact of size, regional lymph nodes metastasis, hypercalcemia, histologic pattern, mitotic count, necrosis, inflammatory and lympho-vascular invasion, anisokaryosis and anisocytosis was also evaluated. Thirty-five dogs were included, twenty-four of which also had metastatic lymph nodes. When the entire population was evaluated, only metastatic disease spread to regional lymph nodes, and necrosis and inflammatory infiltration were correlated to prognosis. When only dogs with metastatic disease were evaluated, size, solid histologic pattern, presence of lymphatic and vascular invasion showed influence on prognosis. Ki67 index was not associated with survival time and disease free interval in any case. The results of this study showed that lymph nodes metastasis at diagnosis reduced disease free interval. Moreover, tumor size greater than 5.25 cm, presence of lymphatic and vascular invasion and a solid histologic pattern were associated with a shorter survival time in dogs with metastasis to regional lymph nodes. Ki67 expression was not significantly associated with prognosis, therefore it could not be considered as a prognostic factor in this tumor type, while the role of hypercalcemia remained unclear.

Benign or Low-Grade Malignant Masses Occupying the Pelvic Canal Space in 11 Dogs.

Dogs with benign intra-pelvic rectal or vaginal masses show symptoms indicating compression on the adjacent organs. Clinical signs usually develop late when the lesion is large enough to interfere functionally. The dogs were referred for severe fecal and/or urinary tenesmus. The data collected included signalment, clinical signs, results of physical examination, pre-surgical diagnostic tests, surgical technique used, surgical complications and histological findings. Digital rectal and vaginal examination allowed the detection of a mass occupying space in the pelvic cavity in all patients. Abdominal ultrasonography and/or total body computed tomography (CT) were used to better characterize the lesion and to exclude a metastatic spread of the tumor in case of malignancy. A dorsal approach to the rectum, a dorsal episiotomy, a midline celiotomy, and a combined perineal and abdominal approach were performed to remove the mass. No postoperative complications were observed. Benign and well-differentiated malignant mesenchymal neoplasms were histologically diagnosed. As a consequence of the chronic urethral compression caused by the mass, urinary incontinence and/or urinary retention were observed for a few postoperative days. Fecal tenesmus resolved in all cases in the immediate postoperative period. The dogs' quality of life quickly improved after surgery, especially considering the serious and life-threatening pre-surgical clinical conditions. Both the recovery time after surgery and overall survival were also evaluated.

Radiofrequency Thermoablation On Ex Vivo Animal Tissues: Changes on Isolated Swine Thyroids.

The use of Radiofrequency thermoablation (RFA) for treating large thyroid nodules is limited by the modest efficiency of the available systems in terms of volume of the ablation zones (AZs). This increases the risk of incomplete ablation of the nodule. Systems employing perfused electrodes have been developed to increase the volume of the AZ. Aim of this study is to compare the size of the AZ induced by RFA systems using internally cooled perfused vs. non-perfused electrodes in swine thyroids. RFAs were performed on 40 freshly isolated swine thyroids using both systems. The perfused system was tested using 0.9% saline, 7% and 18% hypertonic saline solutions. Energy delivery to the tissue was stopped when tissue conductivity dropped (real life simulations) and after an established time of 20 seconds (controlled duration). Following RFA, thyroids were transversally and longitudinally cut. Photographs were taken for macroscopic morphometry of the ablated zones before and after formalin fixation, to evaluate tissue shrinkage. Microscopic morphometry was performed on PAS stained sections. In real life simulation experiments, gross morphometry revealed that AZs produced with electrodes perfused using 7.0% saline are larger compared to isotonic saline. Microscopically, all the conditions tested using the perfused system produced larger AZs compared to non-perfused system after 20 seconds of RFA. In conclusion, the perfusion with 7.0% NaCl solution increased the electrical conductivity of the tissue in real life simulations, resulting in larger ablated areas compared to the use of isotonic saline.

Surgical treatment and outcome of sterile prostatic cysts in dogs.

OBJECTIVE: To describe the surgical treatment and outcome of a large cohort of dogs with sterile prostatic cysts (PCs). STUDY DESIGN: Retrospective study. ANIMALS: Forty-four client-owned dogs. METHODS: Dogs with sterile PCs with at least 6 months of follow-up were included. Clinical variables, type of surgery, complications, recurrences, and outcomes (telephonic interviews or rechecks) were recorded. RESULTS: Extra- and intraparenchymal cysts were diagnosed in 29 and 11 dogs, respectively. Four dogs had both types. Extraparenchymal cysts were treated by partial resection and omentalization (n = 22) and complete resection (n = 7). Drainage and intracapsular omentalization were performed in all dogs with intraparenchymal cysts. The four dogs with both types of cyst were treated by omentalization. Resolution was documented in 39/44 dogs (88.6%). Intraoperative complications occurred in one dog (urethral tear). Major complications resulting in death occurred in three dogs (oliguric kidney injury, cardiac arrhythmia, and persisting urinary tract obstruction). Minor complications (n = 10) consisted of temporary urinary incontinence (n = 2), permanent urinary incontinence (n = 5), urinary retention (n = 2), and dysuria (n = 1). Recurrence occurred in two dogs with extraparenchymal cysts. Median long-term follow-up was 528 days (range, 250-730 days). Thirty-nine dogs had no signs associated with prostatic disease at long-term follow-up. CONCLUSION: Partial or complete resection and/or omentalization of sterile PCs led to resolution of clinical signs in most dogs, although postoperative urinary incontinence was frequent. IMPACT: This study is the largest case series relative to canine sterile PCs treated surgically and provides evidence on the prognosis and rate of complications.

Difference in outcome between curative intent vs marginal excision as a first treatment in dogs with oral malignant melanoma and the impact of adjuvant CSPG4-DNA electrovaccination: A retrospective study on 155 cases.

Canine oral malignant melanoma is locally invasive and highly metastatic. At present, the best option for local control is en bloc excision followed by radiation if excision margins are incomplete. Adjuvantly, the role of chemotherapy is dubious while immunotherapy appears encouraging. This retrospective study evaluated 155 dogs with oral malignant melanomas (24 stage I, 54 stage II, 66 stage III and 11 stage IV) managed in a single institution. The aim was to evaluate the differences in median survival time (MST) and disease-free interval (DFI) between dogs which, at presentation, were treated surgically with a curative intent (group 1) vs those marginally excised only (group 2). MST in group 1 was longer than in group 2 (594 vs 458 days), but no significant difference was found (P = .57); a statistical difference was, however, found for DFI (232 vs 183 days, P = .008). In the subpopulation of vaccinated dogs, the impact of adjuvant anti-CSPG4 DNA electrovaccination was then evaluated (curative intent, group 3, vs marginal, group 4); a significant difference for both MST (1333 vs 470 days, respectively, P = .03) and DFI (324 vs 184 days, respectively, P = .008) was found. Progressive disease was significantly more common in dogs undergoing marginal excision than curative intent excision for both the overall population (P = .03) and the vaccinated dogs (P = .02). This study pointed out that, after staging, wide excision together with adjuvant immunotherapy was an effective approach for canine oral malignant melanoma.

Evaluation of the neoplastic infiltration of the skin overlying canine subcutaneous soft tissue sarcomas: An explorative study.

Studies regarding the neoplastic infiltration of the skin overlying canine subcutaneous soft tissue sarcoma (sSTS) are lacking. In case of the absence of tumor infiltration, there would be the possibility of leaving this unaffected skin in place, thus simplifying surgery. The aim of the study was to investigate whether the skin overlying sSTSs is infiltrated by neoplastic cells. Dogs with sSTSs treated surgically were prospectively enrolled. After excision, the skin was dissected from the tumor along the natural surgical plane of cleavage and histologically evaluated. Twenty-nine dogs with an sSTS were included (22 grade I, 6 grade II, and 1 grade III). The sSTS-overlying skin was not tumor-infiltrated in 14/29 cases (48.3%). A higher frequency of infiltration was observed in higher grade sSTSs (grades II and III, 100%; P = .006); nevertheless, 8/22 grade I sSTSs (36%) also showed cutaneous infiltration. This infiltration involved the dermis of the skin directly in contact with the tumor (multifocal in 11 and diffuse in four cases). Although the cutaneous tumor infiltration is less frequent in grade I sSTSs and a wide excision may still be the safest treatment for any sSTS for a greater possibility of local control, this study opens the possibility to a less aggressive cutaneous excision, but still with a local curative intent, as only the skin directly in contact with the sSTS has been proven to be tumor-infiltrated. Additional studies are warranted to confirm that excision of only this skin may guarantee a complete local control, especially in lower-grade sSTSs.

Evaluation of prognostic impact of pre-treatment neutrophil to lymphocyte and lymphocyte to monocyte ratios in dogs with oral malignant melanoma treated with surgery and adjuvant CSPG4-antigen electrovaccination: an explorative study.

The role of systemic inflammation in cancer's progression has been widely investigated, especially in melanoma in humans. Pre-treatment leukocyte counts and ratios play a recognized prognostic role in several types of malignancies, but no information is available regarding canine oral malignant melanoma (COMM). The purpose of this explorative retrospective study was to investigate the prognostic impact of pre-treatment neutrophil to lymphocyte (NLR) and lymphocyte to monocyte (LMR) ratios in dogs with oral malignant melanoma that underwent surgical resection and immunotherapy with adjuvant CSPG4-antigen electrovaccination. Thirty-nine dogs with histologically confirmed oral melanoma and with available pre-treatment haematological analyses, performed at maximum 60 days before the first treatment, were retrospectively enrolled. Statistical analysis was performed to explore possible correlations among NLR and LMR with age, clinical stage, tumour pigmentation, tumour size, nuclear atypia, mitotic index, Ki67, CSPG4 expression, ulceration, bone invasion and excision margins status. The impact of NLR and LMR on overall survival time (OST) was explored among various ratio cut off and across different time points with Kaplan-Meier method. No significant relationship was identified between leukocytes ratios and histological parameters, CSPG4 expression, excision margin status, age, tumour size and clinical stage. NLR and LMR did not display a prognostic impact on the survival time of the entire population. Pre-treatment leukocyte ratios may not represent a useful prognostic factor in dogs with oral melanoma, especially in absence of distant metastatic disease.

Outcomes of cats treated with maxillectomy: 60 cases. A Veterinary Society of Surgical Oncology retrospective study.

Maxillectomy is poorly described for the management of oral tumours in cats and is occasionally not recommended because of the high complication rate and sub-optimal outcome reported in cats treated with mandibulectomy. The purpose of this study was to retrospectively evaluate the complications and oncologic outcome in cats treated with maxillectomy. Sixty cats were included in the study. Maxillectomy procedures included unilateral rostral (20.0%), bilateral rostral (23.3%), segmental (10.0%), caudal (20.0%) and total unilateral maxillectomy (26.7%). Intra-operative and post-operative complications were reported in 10 (16.7%) and 34 (56.7%) cats, respectively. The most common post-operative complications were hyporexia (20.0%) and incisional dehiscence (20.0%). The median duration of hyporexia was 7 days. Benign tumours were diagnosed in 19 cats (31.7%) and malignant tumours in 41 cats (68.3%). Local recurrence and metastatic rates were 18.3% and 4.9%, respectively; the median progression-free interval (PFI) was not reached. The disease-related median survival time was not reached overall or for either benign or malignant tumours. The 1- and 2-year survival rates were, respectively, 100% and 79% for cats with benign tumours, 89% and 89% for cats with malignant tumours, 94% and 94% for cats with fibrosarcomas, 83% and 83% for cats with squamous cell carcinomas, and 80% and 80% for cats with osteosarcomas. Poor prognostic factors included mitotic index for PFI, adjuvant chemotherapy for both PFI and survival time, and local recurrence for survival time. Maxillectomy is a viable treatment option for cats resulting in good local tumour control and long survival times.

Effect of perioperative desmopressin in cats with mammary carcinoma treated with bilateral mastectomy.

Perioperative administration of desmopressin has shown to significantly decrease rates of local recurrence and metastasis, and increase survival times in dogs with grade II and III mammary carcinomas. The objective of this study was to compare the oncologic outcome of cats with mammary carcinoma treated with bilateral mastectomy with or without perioperative administration of desmopressin. Medical records from nine veterinary institutions were searched to identify cats diagnosed with mammary carcinoma treated with bilateral mastectomy. Sixty cats treated with single-session or staged bilateral mastectomy were included. There were no significant differences in oncologic outcomes found between cats treated and not treated with desmopressin. No adverse effects were seen in any of the cats treated with perioperative desmopressin. Postoperative complications occurred in 18 cats (38.3%) treated with single-session bilateral mastectomy and in three cats (23.1%) treated with staged bilateral mastectomy (P = .48). Histologic grade and a modification of a proposed five-stage histologic staging system were both prognostic for disease-free interval. Incomplete histologic excision was associated with significantly increased rates of metastasis and tumour progression, and a shorter median survival time (MST). Cats that developed local recurrence also had a significantly shorter MST. The results of this study do not support the use of perioperative desmopressin to improve outcome when performing bilateral mastectomy for the treatment of mammary carcinoma in cats.

Characterization of the ablation zones produced by three commercially available systems from a single vendor for radiofrequency thermoablation in an ex vivo swine liver model.

BACKGROUND: Radiofrequency Ablation (RFA) is rarely performed in veterinary medicine. A rationale exists for its use in selected cases of canine liver tumours. RFA induces ablation zones of variable size and geometry depending on the technique used and on the impedance of the targeted organ. OBJECTIVES: (a) to describe the geometry and reproducibility of the ablation zones produced by three commercially available systems from a single company, using isolated swine liver parenchyma as a model for future veterinary applications in vivo; (b) to study the effects of local saline perfusion into the ablated parenchyma through the electrode tip and of single versus double passage of the electrode on size, geometry and reproducibility of the ablation zones produced. METHODS: Size, and geometry of ablation zones reproduced in six livers with one cooled and perfused (saline) and two cooled and non-perfused systems, after single or double passage (n = 6/condition), were assessed macroscopically on digitalized images by a blinded operator. Longitudinal and transverse diameters, equivalent diameter, estimated volume and roundness index were measured. Reproducibility was assessed as coefficient of variation. RESULTS AND CONCLUSIONS: Ablation zone reproducibility was higher when expressed in terms of ablation zone diameters than estimated volume. Local saline perfusion of the parenchyma through the electrode tip during RFA increased the ablation zone longitudinal diameter. Ablation zone estimated volume increased with saline perfusion only when double passage was performed. These data may provide useful information for those clinicians who intend to include RFA as an additive tool in veterinary interventional radiology.

Outcome comparison between radiation therapy and surgery as primary treatment for dogs with periarticular histiocytic sarcoma: An Italian Society of Veterinary Oncology study.

Localized histiocytic sarcoma may occur as a primary lesion in periarticular tissues of large appendicular joints. Treatment options for the primary lesion include radical surgical excision, radiation therapy (RT), or both, in combination with chemotherapy for potential systemic metastases. In an effort to better characterize the time to progression (TTP) following surgical vs non-surgical approaches for periarticular histiocytic sarcoma (PAHS), a contemporary European population of affected dogs was retrospectively surveyed. Medical records were queried for newly-diagnosed PAHS cases undergoing surgery (predominantly limb amputation) or RT followed by systemic chemotherapy. Of 49 dogs, 34 underwent RT and 15 underwent surgery. All dogs received adjuvant chemotherapy. There was no statistically significant difference in TTP or overall survival between groups. The median TTP was 336 days for the operated dogs and 217 days for the irradiated dogs (P = .117). The median overall survival time was 398 days for the operated dogs and 240 days for the irradiated dogs (P = .142). On multi-variable analysis, the variables significantly associated with an increased risk of both tumour progression and tumour-related death were regional lymph node and distant metastasis at admission. Survival and local control rates following RT may be comparable to radical resection. These data may better inform shared decision-making processes between multi-disciplinary care providers and owners.

Caudal Auricular Axial Pattern Flap for the Reconstruction of the Upper Eyelid in Three Cats.

Three cats bearing malignant tumors (two squamous cell carcinomas and one peripheral nerve sheath tumor) affecting the upper eyelid (UE) were treated by en bloc tumor removal. A caudal auricular axial pattern flap was used for reconstruction, and its margin was sutured to only the medial and lateral canthus in two cats; in the third cat, a narrow strip of spared conjunctiva was sutured to the flap margin. Postoperatively, superficial corneal ulcers occurred in two cats, but they healed with topical treatment. Apart from a reduced ability to blink, no further functional deficits persisted, and the long-term cosmetic appearance was considered satisfactory by the owners. Tumor-free excisional margins were achieved in two cases. Tumor recurrence in the long-term was observed for two cats, 350 and 380 days after surgery, one of whom had excisional noninfiltrated margins on histologic examination. UE reconstruction to preserve eyelid function following tumor excision without exenteration is challenging. In selected cases, caudal auricular axial pattern flap is one of the surgical options available to reconstruct the defect resulting from en bloc UE removal without any replacement of the mucosal layer.

In vitro and in vivo effects of toceranib phosphate on canine osteosarcoma cell lines and xenograft orthotopic models.

Canine osteosarcoma (OSA) is the most common primary malignant bone tumour in dogs, and it has a high metastatic rate and poor prognosis. Toceranib phosphate (TOC; Palladia, Zoetis) is a veterinary tyrosine kinase inhibitor that selectively inhibits VEGFR-2, PDGFRs and c-Kit, but its efficacy is not yet fully understood in the treatment of canine OSA. Here, we evaluated the functional effects of TOC on six OSA cell lines by transwell, wound healing and colony formation assays. Subsequently, two cell lines (Wall and Penny) were selected and were inoculated in mice by intrafemoral injection to develop an orthotopic xenograft model of canine OSA. For each cell line, 30 mice were xenografted; half of them were used as controls, and the other half were treated with TOC at 40 mg/kg body weight for 20 days. TOC inhibited cell growth of all cell lines, but reduced invasion and migration was only observed in Penny and Wall cell lines. In mice engrafted with Penny cells and subjected to TOC treatment, decreased tumour growth was observed, and PDGFRs and c-Kit mRNA were downregulated. Immunohistochemical analyses demonstrated a significant reduction of Ki67 staining in treated mice when compared to controls. The results obtained here demonstrate that TOC is able to slightly inhibit cell growth in vitro, while its effect is evident only in a Penny cell xenograft model, in which TOC significantly reduced tumour size and the Ki67 index without modifying apoptosis markers.

Photodynamic Surgery for Feline Injection-Site Sarcoma.

Musculoskeletal sarcomas are rare and aggressive human malignancies affecting bones and soft tissues with severe consequences, in terms of both morbidity and mortality. An innovative technique that combines photodynamic surgery (PDS) and therapy (PDT) with acridine orange has been recently suggested, showing promising results. However, due to the low incidence of sarcoma in humans, this procedure has been attempted only in pilot studies and stronger evidence is needed. Naturally occurring tumors in cats are well-established and advantageous models for human cancers. Feline injection-site sarcoma (FISS) shares with human musculoskeletal sarcomas a mesenchymal origin and an aggressive behavior with a high relapse rate. Furthermore, wide surgical excision is not always possible due to the size and site of development. We assessed the feasibility and the effectiveness of PDS and PDT with acridine orange to prevent FISS recurrence by treating a short case series of cats. For PDS, the surgical field was irrigated with an acridine orange solution and exposed to UV light to enlighten the residual tumor tissue, and the resultant fluorescent areas were trimmed. For PDT, before wound closure, the field was again irrigated with acridine orange solution and exposed to visible light to get the antitumoral cytocidal effect. The procedure was easy to perform and well tolerated, we did not observe any major complications, and all the surgical resection margins were free of disease. Finally, at follow-up, all treated patients did not show evidence of tumor recurrence and had a significantly higher event-free survival rate in respect to a control group treated only by surgery. In conclusion, by this study we demonstrated that, in FISS, PDS and PDT with acridine orange may improve local tumor control, granting a better outcome, and we laid the foundation to validate its effectiveness for the treatment of human musculoskeletal sarcomas.

Identification of CSPG4 as a promising target for translational combinatorial approaches in osteosarcoma.

BACKGROUND: Osteosarcoma (OSA) is a highly metastatic pediatric bone tumor. Adjuvant chemotherapy and surgical resection represent standard treatments; however, the prognosis is still poor. Effective strategies are urgently needed. Chondroitin sulfate proteoglycan (CSPG)4 is a transmembrane proteoglycan with a low expression in normal tissues but high expression in several solid tumors, where it plays a central tumorigenic role. Therefore, it represents a promising therapeutic target. The high homology between human and canine CSPG4 and the recognized translational power of canine tumors as preclinical models for human malignancies prompted us to evaluate CSPG4 expression and the consequences of its immune-targeting for both human and canine OSA treatment. METHODS: We analyzed CSPG4 overexpression in human and canine OSA samples and its significance for the survival of OSA patients. We exploited functional in vitro experiments to assess the antitumor potential of CSPG4 immune-targeting. RESULTS: CSPG4 is overexpressed in OSA and has possible clinical implications as suggested by an evident correlation between CSPG4 overexpression and a shorter survival for both OSA-affected humans and dogs. The potential of CSPG4 immune-targeting for OSA treatment came from the ability of anti-CSPG4 monoclonal antibodies and sera, derived from human-CSPG4-DNA vaccinated canine patients, to significantly inhibit human and canine CSPG4-positive OSA cell proliferation, migration, and osteospheres generation. Moreover, CSPG4 immune-targeting has been shown to potentiate the effect of doxorubicin. CONCLUSIONS: Overall, these results provide the rationale to investigate the CSPG4 immune-targeting as a promising weapon for the treatment of CSPG4-positive OSA canine patients, to be successfully translated to a human setting.