A multi-institutional epidemiologic study evaluating environmental risk factors for feline oral squamous cell carcinoma

Feline oral squamous cell carcinoma (FOSCC) is an aggressive cancer in domestic cats that has no effective treatment option when advanced. Preventative or early diagnostic measures are thus crucial. FOSCC is also a model for human head and neck SCC (HNSCC); strong risk factors in HNSCC include exposure to alcohol, tobacco, areca nut, and high-risk human papillomavirus. Previous studies have identified flea collar and tobacco smoke exposure, feeding canned tuna, canned cat food and cat foods with chemical additives, living in a rural environment, and having outdoor access as risk factors for FOSCC but there was no overlap in the risk factors between studies. In our study, risks for FOSCC were evaluated in an online epidemiologic survey study in 67 cats with FOSCC and 129 control cats. Clumping clay cat litter and flea collar use were significant risk factors for FOSCC on multiple logistic regression with odds ratios of 1.66 (95% CI 1.20–2.30) and 4.48 (95% CI 1.46–13.75) respectively. Crystalline silica is a carcinogen that may be present in all clay cat litters and tetrachlorvinphos is a carcinogen that is present in the most commonly used flea collars in our study. We recommend further investigation into the association between FOSCC and clay-based litter and/or flea collars containing tetrachlorvinphos.     

Assessment of predictive molecular variables in feline oral squamous cell carcinoma treated with stereotactic radiation therapy

This study evaluated molecular characteristics that are potentially prognostic in cats with oral squamous cell carcinoma (SCC) that underwent stereotactic radiation therapy (SRT). Survival time (ST) and progression‐free interval (PFI) were correlated with mitotic index, histopathological grades, Ki67 and epidermal growth factor receptor expressions, tumour microvascular density (MVD), and tumour oxygen tension (pO₂). Median ST and PFI were 106 and 87 days, respectively (n = 20). Overall response rate was 38.5% with rapid improvement of clinical symptoms in many cases. Patients with higher MVD or more keratinized SCC had significantly shorter ST or PFI than patients with lower MVD or less keratinized SCC (P = 0.041 and 0.049, respectively). Females had significantly longer PFI and ST than males (P ≤ 0.016). Acute toxicities were minimal. However, treatment‐related complications such as fractured mandible impacted quality of life. In conclusion, SRT alone should be considered as a palliative treatment. MVD and degree of keratinization may be useful prognostic markers.     

Evaluation of plasma folate and homocysteine concentrations in cats with and without oral squamous cell carcinoma

Feline oral squamous cell carcinoma (SCC) is a devastating disease with an extremely poor long‐term prognosis even with aggressive therapy. Folate and homocysteine derangements are identified in people diagnosed with head and neck SCC. The purpose of this study was to measure plasma folate and homocysteine concentrations in cats diagnosed with oral SCC (n = 13) and to compare these concentrations with those found in cats diagnosed with other tumour types (n = 25), cats with oral, non‐neoplastic disease (n = 6) and healthy cats (n = 24). The median plasma folate concentration in cats diagnosed with oral SCC was 14.7 ng mL^?1, while the median plasma homocysteine concentration was 2.61 μg mL^?1. These concentrations did not differ significantly from those of cats in the other groups. This suggests that different factors may contribute to the pathogenesis of this tumour in cats when compared with people, although evaluation of larger numbers of cats may still identify a difference between groups.     

COMPARISON BETWEEN 2‐18F‐FLUORO‐2‐DEOXY‐D‐GLUCOSE POSITRON EMISSION TOMOGRAPHY AND CONTRAST‐ENHANCED COMPUTED TOMOGRAPHY FOR MEASURING GROSS TUMOR VOLUME IN CATS WITH ORAL SQUAMOUS CELL CARCINOMA

Feline oral squamous cell carcinoma is one of the most refractory feline malignancies. Most patients succumb due to failure in local tumor control. 2‐¹⁸F‐fluoro‐2‐deoxy‐D‐glucose positron emission tomography (¹⁸F‐FDG PET) is increasingly being used for veterinary oncology staging as it highlights areas with higher glucose metabolism. The goal of the current prospective study was to compare gross tumor volume measurements using ¹⁸F‐FDG PET vs. those using computed tomography (CT) for stereotactic radiation therapy planning in cats with oral squamous cell carcinoma. Twelve cats with confirmed oral squamous cell carcinoma underwent pretreatment ¹⁸F‐FDG PET/CT. Gross tumor volumes based on contrast‐enhanced CT and ¹⁸F‐FDG PET were measured and compared among cats. Mean PET gross tumor volume was significantly smaller than mean CT gross tumor volume in the mandibular/maxillary squamous cell carcinoma group (n = 8, P = 0.002) and for the total number of patients (n = 12, P = 0.006), but not in the lingual/laryngeal group (n = 4, P = 0.57). Mismatch fraction analysis revealed that most of the lingual/laryngeal patients had a large region of high‐¹⁸F‐FDG activity outside of the CT gross tumor volume. This mismatch fraction was significantly greater in the lingual/laryngeal group than the mandibular/maxillary group (P = 0.028). The effect of poor spatial resolution of PET imaging was greater when the absolute tumor volume was small. Findings from this study indicated that ¹⁸F‐FDG PET warrants further investigation as a supplemental imaging modality in cats with oral squamous cell carcinoma because it detected regions of possible primary tumor that were not detected on CT images.     

Epidermal growth factor receptor expression in feline oral squamous cell carcinomas

Feline oral squamous cell carcinomas (SCC) have a poor prognosis despite aggressive treatment with surgery, radiation and anticancer drugs. Overexpression of the epidermal growth factor receptor (EGFR), a membrane‐bound tyrosine kinase receptor, has been found in many human epithelial neoplasms, including oral SCC. EGFR overexpression has been associated with advanced disease and a poor prognosis. The purpose of this study was to determine whether feline oral SCC express EGFR. Thirteen formalin‐fixed paraffin wax‐embedded biopsy samples from feline oral SCC were analysed for EGFR expression using immunohistochemistry. Nine of 13 tumours (69%) were positive for EGFR expression, suggesting that altered EGFR expression plays a role in feline oral SCC and provides a rationale for a potential clinical benefit using EGFR inhibitors in combination with conventional treatments.     

EFFECTS OF ANESTHETIC PROTOCOL ON NORMAL CANINE BRAIN UPTAKE OF 18F‐FDG ASSESSED BY PET/CT

The purpose of this study was to assess the effects of four anesthetic protocols on normal canine brain uptake of 2‐deoxy‐2‐[¹⁸F]fluoro‐d ‐glucose (FDG) using positron emission tomography/computed tomography (PET/CT). Five clinically normal beagle dogs were anesthetized with (1) propofol/isoflurane, (2) medetomidine/pentobarbital, (3) xylazine/ketamine, and (4) medetomidine/tiletamine–zolazepam in a randomized cross‐over design. The standard uptake value (SUV) of FDG was obtained in the frontal, parietal, temporal and occipital lobes, cerebellum, brainstem and whole brain, and compared within and between anesthetic protocols using the Friedman test with significance set at P<0.05. Significant differences in SUVs were observed in various part of the brain associated with each anesthetic protocol. The SUV for the frontal and occipital lobes was significantly higher than in the brainstem in all dogs. Dogs receiving medetomidine/tiletamine–zolazepam also had significantly higher whole brain SUVs than the propofol/isoflurane group. We concluded that each anesthetic protocol exerted a different regional brain glucose uptake pattern. As a result, when comparing brain glucose uptake using PET/CT, one should consider the effects of anesthetic protocols on different regions of the glucose uptake in the dog's brain.     

Neoadjuvant chemoradiotherapy and surgery as treatment for oral maxillary squamous cell carcinoma in a dog

A gingival maxillary squamous cell carcinoma was diagnosed in a 12-year-old male Yorkshire Terrier. After a complete diagnostic work-up, including a computed tomography scan, the tumour was staged as T3bN1aM0 and considered non-resectable at presentation. The combination of neoadjuvant megavoltage radiotherapy and neoadjuvant and adjuvant chemotherapy with carboplatin and doxorubicin decreased the size of the tumour, allowing for surgery. The dog was free from local disease for 421 days after which it was euthanased at the owners' request.     

Predicting clinical outcome in feline oral squamous cell carcinoma: tumour initiating cells,telomeres and telomerase

Feline oral squamous cell carcinoma (SCC) has very poor prognosis. Here, a retrospective pilot study was conducted on 20 feline oral SCC patients who underwent stereotactic radiation therapy (SRT), to evaluate: (1) the value of putative tumour initiating cell (TIC) markers of human head and neck SCC (CD44, Bmi‐1); (2) telomere length (TL) specifically in putative TICs; and (3) tumour relative telomerase activity (TA). Significant inverse correlations were found between treatment outcomes and Bmi‐1 expression, supporting the predictive value of Bmi‐1 as a negative prognostic indicator. While TL exhibited a wide range of variability, particularly in very short fractions, many tumours possessed high levels of TA, which correlated with high levels of Bmi‐1, Ki67 and EGFR. Taken together, our results imply that Bmi‐1 and telomerase may represent novel therapeutic targets in feline oral SCC, as their inhibition – in combination with SRT – would be expected to have beneficial treatment outcome.     

Piroxicam and carboplatin as a combination treatment of canine oral non-tonsillar squamous cell carcinoma: a pilot study and a literature review of a canine model of human head and neck squamous cell carcinoma

Results of the treatment with a combination of carboplatin and piroxicam in seven dogs with advanced non‐tonsillar oral squamous cell carcinoma (SCC) were retrospectively analysed. This multi‐agent protocol was well tolerated by all dogs and resulted in a complete regression of the tumour without additional surgery in four of seven patients. Additional surgery was necessary to remove a metastatic lymph node in one dog and residual tumour in a second dog, which achieved a partial response following medical therapy. Median follow‐up for all the dogs was 534 days, while the time‐to‐recurrence, time‐to‐progression and overall survival for this group of patients have not yet been reached. Our study, although limited in number of animals, suggests that this multiagent approach is a useful treatment option for oral non‐tonsillarSCC in dogs and warrants wider application.     

Treatment of canine oral papillary squamous cell carcinoma using definitive‐intent radiation as a monotherapy—a case series

Canine oral papillary squamous cell carcinoma (COPSCC) is a rare neoplasm and although locally invasive it carries a favourable prognosis following wide surgical excision. Radiotherapy has been reported to be effective as an adjunct treatment to surgery. However, limited information is available on the role of radiotherapy as single treatment. This single‐institution retrospective study describes a series of 10 dogs diagnosed with macroscopic COPSCC that were treated with definitive‐intent radiotherapy (DRT) as a monotherapy. These dogs had a median age of 4 years (range: 0.4‐9.6 years). The tumour was located in the rostral oral cavity in all cases with a median tumour size of 2.5 cm (range: 0.8‐6.8 cm). No local or distant metastases were identified. All dogs were treated with electron beam DRT (>32Gy, 10‐16 daily fractions of 3.2Gy). The median follow‐up time was 961 days (range: 333‐3.498 days) with nine dogs achieving a complete response and one dog a partial response. The dog with the partial response developed disease progression at 228 days after initiation of radiotherapy. Two dogs died from non‐tumour‐related causes. The remaining seven dogs were still alive and in complete remission at the time of last follow‐up. Median progression‐free survival time and median survival time were not reached. DRT was generally well tolerated, but all dogs experienced self‐limiting acute radiation mucositis (grade 2‐3) and/or dermatitis (grade 1). No late radiation toxicity was observed. Macroscopic COPSCC appears to be a radiosensitive tumour that can be successfully treated with DRT eliminating the need for aggressive surgery in advanced cases.     

CHARACTERIZATION OF PHYSIOLOGIC 18F‐FDG UPTAKE WITH PET‐CT IN DOGS

We evaluated the whole body distribution of 2‐deoxy‐2‐[18F]fluoro‐d ‐glucose (¹⁸F‐FDG) in seven beagle dogs using positron emission tomography/computed tomography. The mean and maximum standard uptake values (SUV) for various tissues were computed. The SUV of the aortic blood pool was 0.65±0.19. Moderate uptake was present in brain (3.40±1.01). Mild uptake was present in orbital muscles, soft palate, laryngeal and pharyngeal region, mandibular salivary gland, myocardium, liver, pancreas, kidney, and intestine. ¹⁸F‐FDG uptake would be normally higher in these tissues because of normal physiologic activity. Mean and maximum SUV values of the eye, skeletal muscle, bone tissue, spleen, adrenal gland, stomach, tongue, gall bladder, and lung were similar to or lower than that of the aortic blood pool. These data provide a normal baseline for comparing pathologic ¹⁸F‐FDG uptake.     

Environmental risk factors for the development of oral squamous cell carcinoma in cats

BackgroundRisk factors for oral squamous cell carcinoma (OSCC) in cats are derived from a single study dated almost 20 years ago. The relationship between inflammation of oral tissues and OSCC is still unclear.ObjectivesTo investigate previously proposed and novel potential risk factors for OSCC development, including oral inflammatory diseases.AnimalsHundred cats with OSCC, 70 cats with chronic gingivostomatitis (CGS), 63 cats with periodontal disease (PD), and 500 controls.MethodsProspective, observational case‐control study. Cats with OSCC were compared with an age‐matched control sample of client‐owned cats and cats with CGS or PD. Owners of cats completed an anonymous questionnaire including demographic, environmental and lifestyle information.ResultsOn multivariable logistic regression, covariates significantly associated with an increased risk of OSCC were rural environment (OR: 1.77; 95% CI: 1.03‐3.04; P = .04), outdoor access (OR: 1.68; 95% CI: 1.07‐2.63; P = .02), environmental tobacco smoke (OR: 1.77; 95% CI: 1.05‐3; P = .03), and petfood containing chemical additives (OR: 1.98; 95% CI: 1.04‐3.76; P = .04). Risk factors shared with CGS and PD were outdoor access and petfood containing chemical additives, respectively. A history of oral inflammation was reported in 35% of cats with OSCC but did not emerge as a risk factor.Conclusions and Clinical ImportanceThe study proposes novel potential risk factors for OSCC in cats. Although a history of inflammatory oral disease was not significantly more frequent compared with random age‐matched controls, OSCC shared several risk factors with CGS and PD.     

Combined zoledronic acid and meloxicam reduced bone loss and tumour growth in an orthotopic mouse model of bone‐invasive oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is common in cats and humans and invades oral bone. We hypothesized that the cyclooxygenase (COX)‐2 inhibitor, meloxicam, with the bisphosphonate, zoledronic acid (ZOL), would inhibit tumour growth, osteolysis and invasion in feline OSCC xenografts in mice. Human and feline OSCC cell lines expressed COX‐1 and COX‐2 and the SCCF2 cells had increased COX‐2 mRNA expression with bone conditioned medium. Luciferase‐expressing feline SCCF2Luc cells were injected beneath the perimaxillary gingiva and mice were treated with 0.1 mg kg^?1 ZOL twice weekly, 0.3 mg kg^?1 meloxicam daily, combined ZOL and meloxicam, or vehicle. ZOL inhibited osteoclastic bone resorption at the tumour–bone interface. Meloxicam was more effective than ZOL at reducing xenograft growth but did not affect osteoclastic bone resorption. Although a synergistic effect of combined ZOL and meloxicam was not observed, combination therapy was well‐tolerated and may be useful in the clinical management of bone‐invasive feline OSCC.     

Investigation of multiple Felis catus papillomavirus types (-1/-2/-3/-4/-5/-6) DNAs in feline oral squamous cell carcinoma: a multicentric study

Recent evidence suggests a possible association of Felis catus papillomavirus type 2 (FcaPV-2) DNA with feline oral squamous cell carcinoma (FOSCC). In this study, type-specific PCR targeting two genes (L1/E6 or E1/E6) of FcaPV-1/-2/-3/-4/-5/-6 was performed to detect viral DNA in a large amount of FOSCC samples collected in Italy and Austria. FcaPV-1/-2/-3/-4/-5 were detected in 7/113 (6.2%), 7/93 (7.5%), 6/113 (5.3%), 1/113 (0.9%) and 2/113 (1.8%) specimens, respectively, with different prevalences in Italian vs. Austrian samples, whilst FcaPV-6 went undetected. Our results confirms that FcaPV-2 is the most prevalent in FOSCC, followed by FcaPV-1/-3 and suggest that FcaPVs have variable circulation rates in European countries.     

Expression of epidermal growth factor receptor (EGFR) and Ki67 in feline oral squamous cell carcinomas (FOSCC)

The aims of this study were to establish expression of epidermal growth factor receptor (EGFR) and Ki67 in 67 archived biopsy samples of feline oral squamous cell carcinomas (FOSCCs) and to establish if the expression of either markers was predictive of survival. Samples were immunohistochemically labelled for the two proteins and scored. Statistical analyses of data, including Kaplan-Meier survival curves, were performed. All samples expressed both markers although levels differed between samples. Median overall survival was 46 days and 1-year survival was 5%. There was no correlation between Ki67 and EGFR scores (Pearson's correlation coefficient, P = 0.861). Low cellular proliferation (low Ki67 score) was positively correlated with an overall longer survival (Log Rank, P = 0.02) and a trend towards better survival for the high EGFR group was observed (Log Rank, P = 0.076). Ki67 and EGFR immunostaining in FOSCC may be of value as biochemical markers for screening of biopsies from cases of FOSCC.     

Electrochemotherapy in treatment of canine oral non‐tonsillar squamous cell carcinoma. A case series report

Non‐tonsillar squamous cell carcinoma (ntSCC) is a common and locally aggressive oral tumour in dogs. The treatments of choice are currently surgery and radiotherapy. Electrochemotherapy (ECT) is a local ablative anti‐tumour technique using electric pulses to enhance the intracellular diffusion of cytotoxic drugs. The aim was to retrospectively evaluate the outcome of patients with oral ntSCC treated with ECT. Twelve dogs with ntSCC were retrospectively enrolled. ECT was combined with IV bleomycin (15 000 UI/m²) alone in 11 cases and post‐surgery in 1. Parameters considered were: tumour site and size, electroporation parameters, response rate (complete remission [CR], partial remission [PR]), median survival time (MST), recurrence rate (RR), median disease‐free interval (DFI) and treatment toxicity (6‐point scale). Median tumour size was 1.65 cm (range 0.3‐8.0 cm) and the response rate was 90.9% (10/11; 8 CR and 2 PR). Two dogs underwent a second ECT. MST for dogs dead with tumour (n = 2) was 110 days and for dogs dead without tumour (n = 3) was 831 days. Among five surviving dogs, one experienced tumour recurrence and four were in CR. Results from two dogs were analysed separately. Overall RR was 27.3%. DFI and MST for dogs with recurrence were 50 and 115 days, respectively. Treatment toxicity was very low. We noticed that all dogs with tumours smaller than 1‐2 cm achieved CR without recurrence suggesting a favourable prognosis when using ECT. ECT for canine ntSCC could be considered a valid treatment option especially for smaller tumours, but a larger caseload would be needed to confirm this statement.     

The expression of calretinin and cytokeratins in canine acanthomatous ameloblastoma and oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) and canine acanthomatous ameloblastoma (CAA) represent two epithelium‐derived neoplasms that affect the oral cavity of dogs. The expression of cytokeratins (CKs) and calretinin has been previously established in the canine tooth bud and odontogenic tumours. The aim of this study was to characterize the CK and calretinin expression profile of OSCC in comparison to CAA and canine tooth bud tissues. Samples from 15 OSCC and 15 CAA cases, as well as 6 tooth buds and 2 normal gingival tissues were examined. OSCC CK expression was consistent with the CK expression profile of CAA and canine tooth bud tissue. Calretinin was positively expressed in 10 of 15 OSCC cases, with 5 cases demonstrating high staining intensity. Only 2 of 15 CAA cases demonstrated mild‐moderate staining intensity. The statistically significant difference in staining pattern and intensity of calretinin in OSCC and CAA can help distinguish between these two tumour types.     

Detection of papillomaviral DNA sequences in a feline oral squamous cell carcinoma

Oral squamous cell carcinomas (OSCCs) are common and often fatal feline neoplasms. Factors that predispose to neoplasm development in cats are poorly defined. Around 25% of human OSCCs are caused by papillomaviruses (PVs). To determine if PVs are associated with OSCCs in cats, three sets of consensus primers were used to evaluate 20 feline OSCCs and 20 non-neoplastic feline oral lesions for the presence of PV DNA. Papillomaviral sequences were detected within one OSCC, but no non-neoplastic lesion. Sequencing of the amplified DNA revealed a previously unreported PV that was most similar to human PV type 76. This is the first time PV DNA has been amplified from the oral cavity of a cat. However, while these results suggest that feline gingival epithelial cells can be infected by PVs, they do not support a causal association between viral infection and the development of feline OSCCs.