Non-ocular melanomas in cats: a retrospective study of 30 cases

Abstract

Objectives

The aim of the study was to describe the clinical outcome of 30 cats with non-ocular melanomas and to evaluate the association between clinical or pathological parameters and overall survival time.

Methods

The database of the animal histopathological laboratory of the National Veterinary School of Nantes (Oniris, Nantes, France) was retrospectively searched to identify cases of feline non-ocular melanomas between December 2009 and April 2014. For each case, clinical data, including signalment, location of the primary tumour, staging, treatment and outcome, were collected from the medical records or via interviews with referring veterinarians. Histological and immunohistochemical evaluation included mitotic index, cytonuclear atypias, junctional activity, Melan A and S100 immunostaining, and surgical margins. Univariate analysis to test the prognostic value of the different variables was performed by the Kaplan–Meier product limit method using the log-rank test of significance.

Results

Thirty cats were included in the study. Eleven had a cutaneous non-auricular melanoma, six had a tumour located on the pinna and 13 had a tumour in the oral cavity. Cats with auricular melanomas were significantly younger than cats with tumours in other locations. Location and presence of clinical signs were not of prognostic significance, but the achromic phenotype was significantly associated with a poorer prognosis. Twenty cats were treated with surgery and survived significantly longer than cats that received only medical treatment or that did not receive any treatment. According to our data, mitotic index, cytonuclear atypias, junctional activity, Melan A or S100 expression, and surgical margins were not associated with survival.

Conclusions and relevance

We show for the first time, in a large series, that the auricular form of melanoma affected significantly younger cats than other extraocular forms. Most feline non-ocular melanomas are malignant and achromic tumours are associated with a poorer prognosis. According to this study, surgery should be considered as a priority.

Introduction

Melanomas are tumours derived from melanin-synthesising cells called melanocytes or their precursors, melanoblasts. ¹ Melanocytic neoplasms are common in both dogs and humans. The oral cavity is the most frequent site of malignant melanomas in dogs and it usually carries a poor prognosis because of a high propensity to local invasiveness and a high metastatic rate.^2–6 The cutaneous tissue is the preferential location of melanomas in humans. Ultraviolet (UV) light exposure is a well established promoting factor for cutaneous melanomas, currently the cancer with the highest increase in incidence in people. ⁷

Unlike canine species and human beings, melanocytic neoplasms are rare in cats.^8–13 The eye is the most commonly involved site and ocular melanomas are the most frequent intraocular tumours in cats. ¹⁰ Few data are available concerning non-ocular melanomas. As a consequence, the biological behaviour of these tumours remains relatively unknown. There are only a few studies in which the clinical evolution of cats with non-ocular melanomas is reported. Furthermore, these studies had conflicting results.^3,11

The aim of this study was to describe the clinical aspects of this disease in a series of cats with a diagnosis of non-ocular melanoma and to compare our data with previous studies, to contribute to a better understanding of feline non-ocular melanocytic tumours.

Material and methods

The database of the animal histopathology laboratory of the National Veterinary School of Nantes (Oniris, Nantes, France) was searched to identify all cases with a histopathological diagnosis of non-ocular melanoma from December 2009 to April 2014. Data were collected by reviewing medical records or by telephone interview with or via a pre-established questionnaire from the referring veterinarian. For all cases, signalment (age, sex, breed), location of the primary mass or nodule, the presence of clinical signs, the size and the colour of the primary lesion, the results of staging procedures at diagnosis, intended treatments and outcome were recorded. All cases lost to follow-up after diagnosis were excluded from the study. Histopathological reports were reviewed and data regarding mitotic index, frequency of cytonuclear atypias, junctional activity (ie, the presence of aggregations of melanocytes along the dermoepidermal junction) and surgical margins were gathered. Immunohistochemical (IHC) evaluation of the Melan A and S100 antigen were performed, as previously described, to confirm the melanocytic nature of the tumour. ¹⁴ An automated IHC (Benchmark XT Ventana; Roche Diagnostics) was used with a cross-reactive monoclonal mouse anti-human Melan A antibody (clone A103, dilution 1:25; Novocastra) and a polyclonal rabbit anti-S100 antibody (Z0311, dilution 1:500; Dako). After heat-induced antigen retrieval (60 mins, 95°C), a 32 min incubation time of the primary antibody was applied and fixation was detected by the I-view universal DAB detection kit (Roche Diagnostics), using a polyclonal goat anti-mouse IgG antibody (E433, dilution 1:300; Dako) for Melan A and a polyclonal goat anti-rabbit IgG antibody (E432, dilution 1:300; Dako) for S100.

Univariate analyses to assess the prognostic value of the different variables were performed using the Kaplan–Meier product limit method with the log-rank test of significance. Overall survival was defined as the time between histopathological diagnosis and death. Cats still alive at the end of the study were censored from the survival analysis. The Student’s t-test was performed to compare the distribution of quantitative parameters (eg, age) and Fisher’s exact test was used to compare the distribution of qualitative parameters. All analyses were performed with R statistical software (http://www.r-project.org). Values of P ⩽0.05 were considered significant.

Results

Thirty-nine cases were identified from the digital database. Nine cases were excluded because of loss to follow-up or because reviews of the original slides were not clearly consistent with a melanocytic neoplasm. Thirty cases were finally included in the study: six cats (20.0%) had a tumour located on the pinna, 11 cats (36.7%) had a cutaneous non-auricular melanoma and 13 (43.0%) cats had an oral melanoma. The study population, as detailed in Table 1 in the supplementary material, was composed of 12 neutered males (40.0%), nine spayed females (30%), five intact males (16.7%) and four intact females (13.3%). There were 26 domestic shorthair cats (86.7%), three Siamese (10.0%) and one Ragdoll (3.3%).

Clinical data concerning the first group of six cats with an auricular melanoma are summarised in Table 2 in the supplementary material. There were three neutered males (50%), two spayed females (33.3%) and one intact female (16.7%). The male to female ratio for this group of animals was 1:1. The mean age was 7.9 years (range 4.0–16.0 years). Cats with auricular melanomas were significantly younger than cats with melanomas in other locations (P = 0.036; Figure 1). The lesions were unilateral in five cases and bilateral in one case. Three cats had tumours located at the extremity of the pinna and one at the base. For the two remaining cats, tumour location was not reported. Pruritus was only reported in one cat (cat 1) and clinical signs compatible with inflammation were described in two (cats 1 and 6). Two cats had an evaluation of the regional lymph node on physical examination for which no abnormality was noted. Two cats had thoracic radiographs that did not show any sign compatible with metastasis. As depicted in Table 3 in the supplementary material, five lesions were histologically classified as aggressive (moderate to high mitotic index and/or cytonuclear atypias considered to be moderate to frequent) and one as benign (low mitotic index and rare-to-moderate cytonuclear atypias). Junctional activity was described in three samples. In two cases, tumours were amelanotic. Both were positive for S100, but only one was positive for Melan A at IHC analysis. All cats were treated by partial or complete pinnectomy; two of them had local recurrence. Evaluation of margins was possible in five cases; for the sixth case, the sample sent to the laboratory did not allow for an evaluation of margins. In one case, there was evidence of an infiltration of the margins by the tumour cells, the other four cases were considered to have tumour-free margins. Three cats were still alive, including the cat with a histologically benign lesion, with a follow-up of at least 10 months. As a consequence the median survival time was not reached for this group (Figure 2).

Figure 1.

Box and whisker plots depicting the distribution of the cats’ age according to tumour location. Median age was 7.9 years for cats with auricular tumours, 13.3 years for cats with cutaneous non-auricular tumours and 13.0 years for cats with oral tumours. Cats with auricular tumours were significantly younger than cats with a tumour in another location (P = 0.0359)

Figure 2.

Kaplan–Meier survival curves of cats with auricular, cutaneous non-auricular and oral tumours. At the time of writing, median survival time for cats with auricular melanomas was not reached. Median survival time for cats with cutaneous non-auricular and oral tumours was, respectively, 179 and 120 days. The difference of survival between groups was not significant

The second group of 11 cats had a non-auricular cutaneous melanoma (Table 4 in the supplementary material). There were four neutered males (36.4%), four spayed females (36.4%) and three intact females (27.2%). The male to female ratio for this group of animals was 1:1.75. The mean age was 13.3 years (range 9.0–19.0 years). Six cats had tumours located on the head, two on the trunk and one on the footpad. Tumour location was not reported for the two other cases. Pruritus was only reported in one cat (cat 7) and clinical signs compatible with local inflammation occurred in four (cats 7, 9, 15 and 16). Two cats had a manual evaluation of the regional lymph node and a lymphadenopathy was reported in one of them. Neither histological nor cytological examination of this abnormal lymph node was performed. Histological analysis of the draining lymph node was performed in one cat and revealed a metastatic infiltration. Four cats had thoracic radiographs available. In two cases no sign of metastasis was described, in one case a multinodular opacification was reported and in one case abnormalities concerning the pulmonary parenchyma were described without further precision. Ten lesions were histologically classified as aggressive and one as benign (Table 3 in the supplementary material). Junctional activity was present in six samples. Five tumours were achromic, and all were negative for Melan A but positive for S100. Six cats were treated by surgery alone, two with glucocorticoids alone, one with surgery and corticoids, one received neither medical nor surgical treatment, and the presence of any treatment was not reported in the last cat. Two of the seven cats that had surgical excision of their tumour experienced local recurrence. Evaluation of margins was available in six cases. There was evidence of margins infiltration in three cases; two of them had local recurrence. The other three cases were considered free. Two cats were still alive at the end of the study with a minimum follow-up of 11 months and two died from causes unrelated to the tumour. Median survival time for this group was 179 days (95% confidence interval [CI] 92 days [upper limit of the interval could not be calculated]; Figure 2).

The third group was composed of 13 cats with an oral melanoma (see Table 5 in the supplementary material). There were five neutered males (38.5%), five intact males (38.5%) and three spayed females (23.0%). The male to female ratio for this group was 3.3:1 and was not significantly different from the other groups. The mean age was 13 years (range 10.0–15.0 years). Five tumours were located on the lip, three on the gingiva and the precise location in the oral cavity was not reported for the last three cases. Seven cats had clinical manifestation related to the tumour, six were anorexic and one was anorexic with ptyalism. The presence of clinical signs was significantly more frequent in this group (P = 0.01959). Two cats had manual evaluation of the regional lymph node, and a lymphadenopathy was reported in one of them. Cytological evaluation of this abnormal lymph node was not performed. Three cats had thoracic radiographs available; none had signs of metastasis. All tumours were histologically classified as aggressive (Table 3 in the supplementary material). Junctional activity was present in five samples. Ten lesions out of 13 were achromic; 9/10 stained positively for S100 and 4/10 for Melan A. The frequency of amelanotic tumours among oral melanomas was not significantly different from the proportion in other locations (P = 0.07). Five cats were treated only with surgery, two with glucocorticoids only, two cats had surgery and glucocorticoids, one received non-steroidal anti-inflammatory drugs, one received masitinib and the two latter cats did not receive any treatment. Among the seven cats treated with surgery, five had a local recurrence. Evaluation of margins was available in five cases and marginal infiltration was observed in three cases that all presented local recurrence. Two cats were still alive at the end of the study, with a minimum follow-up of 11 months. Median survival time for this group was 120 days (95% CI 58 days [upper limit of the interval could not be calculated]; Figure 2).

There was no significant difference between the three groups regarding overall survival (P = 0.236; Figure 2). Mitotic index, presence of a junctional activity, infiltration of margins, cytonuclear atypias, presence of clinical signs and presence of inflammation were not associated with survival. Lymph node and/or pulmonary metastasis were assessed in 13 cases and the presence of metastasis was suspected in four, so the metastatic rate was evaluated at 30.7%. Cats treated by surgery survived significantly longer than others (P = 0.009; Figure 3). The absence of pigmentation was significantly associated with a shorter survival (P = 0.0004; Figure 4). Among the 17 amelanotic melanomas, five were immunopositive for Melan A and 16 were immunopositive for S100. In this study, neither Melan A nor S100 expression were associated with survival.

Figure 3.

Kaplan–Meier survival curves of cats treated with or without surgery. Median survival time for cats that were treated by surgery was 143 days, whereas others had a median survival time of 71 days. Cats that underwent surgery lived significantly longer (P = 0.0087)

Figure 4.

Kaplan–Meier survival curves of cats with pigmented or amelanotic tumours. Median survival time for cats that had pigmented tumours was 179 days, whereas others had a median survival time of 71 days. Pigmentation was significantly associated with prognosis (P = 0.0004)

Discussion

Non-ocular melanomas are considered rare in cats, representing only 0.8–7.0% of cutaneous neoplasms and <1% of oral tumours.^15,16 Therefore, the aim of this study was to describe the clinical evolution in a relatively large series of cases in order to improve our understanding of this disease.

The first striking observation was that, even if melanoma tends to be a disease of older individuals, cats with auricular melanomas were significantly younger than the others. In the study of Luna et al, ¹¹ 2/3 cats with auricular melanoma were younger than 10 years of age. Similarly in the study of Miller et al, ¹⁴ 9/16 cats with auricular melanomas were younger than 10 years of age, but in the latter study median age of the total panel was relatively low. Our study establishes, for the first time, a significant difference between the ages of these melanoma patient groups.

In the present study, more than half of the cutaneous melanomas were located on the head. The high frequency of non-ocular melanomas on the cats’ head or ears is consistent with previous studies.^10,11,15 This distribution looks like that of squamous cell carcinomas, which are known to be related to UV light. ¹⁷ Furthermore, in our study one cat had a bilateral auricular tumour. Together, these observations suggest that UV light may be implicated in the development of melanomas in that location. In people, the role of UV radiation in the pathogenesis of cutaneous melanomas is well recognised. In cats, a relationship between non-ocular melanomas and exposure to sunlight has already been suggested, but has never been proven.^11,18 Unfortunately, in this study, the colour of the skin or the coat around the tumour bed was not recorded in the large majority of cases. Thus, an association between pigmentation and development of a melanoma could not been evaluated.

In our study, IHC was performed in amelanotic tumours, to confirm their melanocytic origin. We showed that 12/17 (70.6%) of amelanotic melanomas were negative for Melan A. This is in accordance with a previous study, which reported that 75% of feline amelanotic melanomas were negative for Melan A, and confirms the poor sensitivity of this marker. ¹⁴ On the contrary, almost all the amelanotic melanomas stained positively for S100. This marker is considered as a sensitive marker for melanocytic neoplasms but with a low specificity, as other tumours (eg, peripheral nerve sheath tumours, chondrosarcomas) may also stain positive for this marker. In the feline species, there is no other validated marker to confirm the melanocytic nature of a tumour. Therefore, in this study, even in cases of negative Melan A immunostaining, the presence of positive immunostaining for S100 in the context of a compatible histopathological morphology was considered sufficient to confirm the diagnosis of a melanocytic neoplasm.

In this study, amelanotic lesions were associated with a poorer prognosis. A previous study showed that amelanotic melanomas were associated with the histological criteria of malignancy, which correlated with an aggressive biological behaviour in 93.3%. ¹⁵ Amelanotic melanomas in dogs also have a poorer prognosis. ¹⁹ The absence of pigments in the tumour cells could be related to dedifferentiation and to the acquisition of a molecular phenotype associated with a higher ability for local invasion and metastatic spread.

Cats that had surgery survived significantly longer than those that received medical treatment only (corticosteroids in the majority of cases). In dogs, wide surgical resection remains the best therapeutic modality to achieve local control, whatever the location of the tumour. ¹⁷ Even if some authors suggested that it could be the same in cats, it has never been proven. ¹¹ Surprisingly, free margins were not associated with a longer survival. This can be due to the relatively low number of cases or to the fact that most surgeries were performed in first-opinion practices that do not carry out procedures associated with carcinogenesis. In fact, tumour samples that were sent to the laboratory might not be representative of the whole tumour and its margins. As an example, for canine oral melanomas, up to 57% of tumours invade bones, necessitating partial or complete maxillectomy or mandibulectomy, which are rarely performed by general practitioners. ¹⁷ The high local recurrence rate (45%) observed in this study could reflect the need for a more aggressive surgical procedure, especially for oral tumours. Moreover, some cases evaluated as having ‘free margins’ might have already been metastatic, thereby having a shorter overall survival.

Metastatic rates have been previously reported to range between 5% and 25% in feline non-ocular melanomas.^10,18 Our data suggest a slightly higher rate, around 30%. However, only 13 cats were evaluated for the presence of local or distant metastasis and this evaluation was partial in most cases (only one cat had histological evaluation of the draining lymph node). This observation underlines the need for a complete staging to evaluate correctly the metastatic rate of any neoplastic process.

With regard to canine melanomas, radiation therapy is currently the recommended treatment after surgery or in a palliative setting. The overall response of canine oral melanomas to radiation therapy ranges from 82% to 94%. ¹⁷ One study evaluated the efficacy of a hypofractionated radiation therapy protocol in five cats with oral melanomas. One complete response and two partial responses were observed, but all cats in this study were euthanased because of progression of the disease. ⁸ The efficacy of radiation therapy in non-ocular melanomas needs to be better characterised, but it seems that it could be an interesting adjuvant therapy to surgery. Given the metastatic potential of non-ocular melanomas in cats, there could be some rationale for the use of chemotherapy in order to control the metastatic process. However, the efficacy of such therapy in that precise indication has never been evaluated in cats and is controversial in dogs. Finally, a therapeutic DNA vaccine encoding the human tyrosinase has been licensed for the treatment of melanomas in dogs and may be interesting in cats as well, but there is today no report concerning the use of this vaccine in cats.^20,21

A limitation of our study was its retrospective setting: staging procedures, treatment plans and follow-up schedules were not standardised, making comparisons difficult. Despite this, all the cases benefited from a complete pathological evaluation and a complete follow-up in order to determine the life status at the end of the study, as well as the probable cause of death, related or not to the cancer. Almost all cases were treated in first-opinion practices by a wide range of practitioners with different levels of experience. This could have had an impact on diagnostic and staging procedures and also on therapeutic intervention. This is particularly true regarding surgical treatment, the efficacy of which is linked to the surgeon’s experience. However, the study does reflect the current outcome of most cats with this disease.

Conclusions

Non-ocular melanomas are mainly located on the head, on the pinnae and in the oral cavity. Auricular tumours seem to appear in younger cats and an amelanotic phenotype correlates with a worse prognosis. Surgery is of clinical benefit and wide surgical excision of tumours should be recommended first for local control.