An 8-year-old grey Quarter Horse gelding was referred for evaluation of a rapidly growing mass associated with the third eyelid of the left eye. A pigmented mass of approximately 2 cm in diameter was palpated and visualised associated with the conjunctival lining of the nictitans. It was not possible to palpate normal nictitans deep to the base of the mass. A full dermatological examination revealed no other melanomas in common sites. Based on the size and rapid growth of the mass, surgical excision and one application of local chemotherapy was performed under general anaesthesia. Histopathology confirmed the diagnosis of malignant melanoma and the presence of clean surgical margins. There was no recurrence at 5 weeks post surgery. To our knowledge, this is the first report of a primary malignant melanoma of the third eyelid in a horse.
Melanomas are common cutaneous tumours in horses, accounting for 6–15% of all reported skin tumours in this species (Pulley and Stannard 1990). Melanomas occur frequently in older grey horses where a prevalence of 80% has previously been reported (McFadyean 1933), especially in the perineal region, base of the tail, lips and eyelids (Pulley and Stannard 1990; Fleury et al. 2000). Neoplasia of the third eyelid is common in horses, with squamous cell carcinoma most frequently reported (Lavach and Severin 1977; Sundberg et al. 1977; Gearhart et al. 2007; Mathes et al. 2011). A pigmented squamous cell carcinoma has been previously described in a horse in an ocular location (McCowan and Stanley 2004). Other tumours affecting the nictitans include lymphangiosarcoma (Puff et al. 2008), lymphangioma (Gehlen and Wohlsein 2000), papilloma (Lavach and Severin 1977; Junge et al. 1984), haemangiosarcoma (Sansom et al. 2006; Gearhart et al. 2007; Wegge et al. 2009), basal cell tumour (Baril 1973), solid carcinoma (van der Linde-Sipman et al. 1986), sebaceous adenocarcinoma (Kunze and Tvedten 1979) and lacrimal adenocarcinoma (Mathes et al. 2011).
Squamous cell carcinoma is commonly treated by surgical excision with or without adjunctive treatments such as cryotherapy, hyperthermia, radiation therapy or intra-lesional chemotherapeutic agents (Giuliano 2010).
To our knowledge, this is the first report of a malignant melanoma in the nictitans in a horse.
An 8-year-old grey Quarter Horse was presented for evaluation and treatment of a suspected melanoma on the third eyelid of the left eye. The owner noticed mucoid discharge from the left eye and a mass associated with the medial canthus 2 weeks prior to presentation. The horse was in regular exercise and no other health problems were present. After performing an ophthalmic examination, the local veterinarian suspected a melanoma involving the third eyelid and referred the horse for further examination and possible surgical treatment.
On presentation, epiphora was noted at the left eye. An auriculopalpebral nerve block was performed using 3 ml lidocaine 2% (Lidocaine)1. Topical proparacaine (Alcaine)2 was applied to the cornea. Upon digital palpation of the nictitating membrane, a firm, nodular mass, approximately 2 cm in diameter, could be palpated. The ventral margin of the mass was not palpable. The nictitating membrane was subsequently grasped and elevated with Graefe fixation forceps to examine its posterior surface. The mass was pigmented and involved the leading edge of the nictitating membrane and its conjunctival lining on the posterior surface. Full ophthalmic examination of the left eye revealed a moderate conjunctivitis and no fluorescein uptake associated with the mass rubbing on the surface of the cornea. The cornea, anterior chamber and iris were found to be normal. The right eye was also examined and no abnormalities noted. Differential diagnoses for the observed mass were neoplasia (squamous cell carcinoma, pigmented squamous cell carcinoma, lymphangioma/lymphangiosarcoma, papilloma, haemangiosarcoma, basal cell tumour, sebaceous or lacrimal adenocarcinoma and melanoma), granuloma or parasitism (habronemiasis, onchocerciasis), with melanoma being the highest on the list due to the black pigmentation of the mass and its rapid growth.
A full dermatological examination revealed no other melanomas in common sites. Surgical excision and histopathological examination of the mass were recommended.
Prior to surgery, the horse was treated with procaine penicillin G (PenOne Pro)3 22,000 iu/kg bwt i.m., flunixin meglumine (Banamine)4 1.1 mg/kg bwt i.v. and tetanus toxoid4 i.m. The horse was sedated with xylazine (Anased)5 1 mg/kg bwt i.v and butorphanol (Torbugesic)6 0.01 mg/kg bwt i.v. prior to induction with ketamine (Ketaset)6 2.2 mg/kg bwt i.v and diazepam7 0.05 mg/kg bwt i.v and maintainance with inhalational isoflurane. The horse was placed in lateral recumbency with the affected eye uppermost.
The area was routinely prepared for surgery using diluted povidone-iodine solution. The cornea was subsequently lavaged with sterile saline.
The third eyelid was exteriorised and 2 curved Kelly forceps were clamped across the base of the nictitans membrane below the mass to isolate it. The Kelly forceps formed a ‘V’, with the tips at the base of the eyelid and involved all the cartilage. Once isolated, the mass was determined to be entirely removable via excision of the third eyelid. The mass and affected third eyelid were resected using a No. 10 surgical blade, incising the posterior surface first to avoid iatrogenic damage to the cornea. Surgical margins were marked with China ink and the mass submitted for histopathology. As adjunct therapy, the margins of the third eyelid remnant were injected with 10 mg/ml carboplatin in sesame oil, at a dose of 1.5 mg/cm3 as previously described for intra-lesional use (Théon et al. 1996). Preparation, administration and handling of waste material and excreta followed the guidelines for safe use of cytotoxic agents. Triple antimicrobial ophthalmic ointment was applied to the cornea. The horse recovered uneventfully from anaesthesia and was discharged the same day.
The owner was instructed to continue the topical triple antimicrobial for 7 days and to administer flunixin meglumine (Banamine paste)8 0.5 mg/kg bwt orally twice a day for 3 days.
Histologically, the mass was located in the conjunctival submucosa. The mass was unencapsulated and densely cellular. The neoplastic cells were arranged in sheets and had distinct cytoplasmic borders and moderate to abundant amounts of cytoplasm that contained numerous, variably sized, dark brown to black granular pigment consistent with melanin (Fig1). Bleaching of tissue sections removed the dark pigment allowing better evaluation of cellular and nuclear detail (Fig2). There was moderate variability in cellular size and shape. Nuclei were round to pleomorphic with finely stippled chromatin and 1–3 prominent nucleoli. There were 3 mitotic figures counted in 10 randomly selected 400x fields of view. Histopathology confirmed a diagnosis of malignant melanoma of the third eyelid based on cellular and nuclear pleomorphism, increased mitotic index and invasion of the neoplastic cells into surrounding dermal collagen. There was no extension of lesion to the margins of the resected tissue and complete excision was confirmed.
Five weeks post surgery, the surgical site appeared healed with no gross evidence of recurrence.
During embryonic development, melanocyte precursors (melanoblasts) migrate to the skin, eye, meninges, intima of the heart and blood vessels, adrenal glands and autonomic nervous system (Pulley and Stannard 1990). Melanocytes can potentially undergo neoplastic transformation in any of those sites (Kirker-Head et al. 1985), generating a primary melanoma.
The most common clinical presentations in horses have been classified as discrete dermal melanoma and dermal melanomatosis (Valentine 1995). Their histological appearance is similar and is not predictive of the potential for metastases or behaviour (Valentine 1995; Johnson 1998).
Currently, no definitive diagnostic test for differentiation of benign and malignant melanocytic neoplasms or prediction of survival time is available (Giuliano 2010). Usually, cutaneous melanomas grow slowly for up to 20 years without metastasising. Less frequently, they will grow slowly and then undergo malignant transformation. A minority will show rapid growth and malignancy from the onset (Patterson-Kane and Ginn 2003). Malignant melanomas can metastasise anywhere in the horse (Valentine 1995). Reported sites of metastasis include regional or distant lymph nodes or both, blood vessels including pulmonary alveolar capillaries, spleen, liver, mesentery, serosal surface of the intestines, omentum, mediastinum, cranial vault, bone and bone marrow, skeletal muscle and parotid salivary gland. Less commonly, metastases are found in the adrenal glands, kidney, diaphragm, heart, guttural pouches and mammary gland (MacGillivray et al. 2002).
Primary nondermal melanomas are uncommon in horses (Pulley and Stannard 1990). They have been described in the guttural pouch (Sasaki et al. 2007), parotid region (MacGillivray et al. 2002), orbit (Runnels and Benbrook 1941; Seltenhammer et al. 2003), chest, abdomen (Milne 1986; Murray et al. 1997; Tarrant et al. 2001) and epidural space (Traver et al. 1977).
Large melanocytic masses can cause mechanical problems as they obstruct the anal sphincter, prepucial opening or the vulvar commissure: in these cases, treatment is necessary (Johnson 1998). Excision of melanocytic nevi and dermal melanomas has been reported to be curative (Valentine 1995; Johnson 1998). More recently, surgical excision was found to be a viable option also for dermal melanomatosis (Rowe and Sullins 2004).
Reports of treatment of adnexal melanomas in horses are few (Moore et al. 2000; Giuliano 2010). Discrete melanomas of the eyelids are common, slow growing, localised and mostly benign; therefore can be successfully excised (McFadyean 1933; Blodi and Ramsey 1967; Dugan 1992; Moore et al. 2000; Giuliano 2010). Similarly, epibulbar or limbal melanomas are slow growing and respond well to excision and local therapy (Hirst et al. 1983; Hamor et al. 1997). However, conjunctival melanomas can be aggressive in man (Jacobiac et al. 1989) and domestic species (Cook et al. 1985; Collins et al. 1994), including horses (Moore et al. 2000).
In this case, the mass appeared to be rapidly growing, and if confirmed as a melanoma histologically, was in an unusual location. Therefore, aggressive treatment was considered appropriate. One study reported successful induction of tumour regression in extra-ocular melanomas treated with cimetidine at the dose of 2.5 mg/kg bwt orally every 8 h (Goetz et al. 1990); however, subsequent studies evaluating the use if cimetidine have found it to be ineffective (Bowers et al. 1994; Warnick et al. 1995; Laus et al. 2010). There are no studies on the use of cimetidine for adnexal melanomas; it was, therefore, not considered as an appropriate treatment in the present case.
McCauley et al. (2002) described the use of a CO2 laser to ablate nonocular melanomas. Surgical resection and local photodynamic therapy have been successfully combined in one case of a lower eyelid melanoma. No recurrence was noted at 5 years post treatment (Giuliano et al. 2005). The option of using a CO2 laser in this case was excluded due to the vicinity of the lesion to the cornea and the potential for damaging it.
Although not administered in this case, cryotherapy is also believed to have an effect on melanocytic tumours (Jacobiac et al. 1980; Seregard 1998; MacGillivray et al. 2002).
Electrochemotherapy, a combination of intralesional chemotherapy, namely cisplatin and delivery of electric pulses to increase the efficacy and uptake of the drug, has been reported to be successful in decreasing the size of extensive labial melanomas in a horse (Spugnini et al. 2011) and could be considered provided the availability of the necessary equipment.
Immunotherapy offers a more targeted and therefore precise approach to the treatment of cancer. Tyrosinase expression appears constitutively increased in all malignant melanocytic tumours (Ferguson and Kidson 1997; Vourc'h-Jourdain et al. 2009); therefore, tyrosinase has been identified as a potential target for immunotherapy and has been successfully used for this purpose in mice (Ferguson et al. 2008).
A xenogeneic DNA vaccine targeting tyrosinase has been developed and successfully used in dogs with melanoma (Bergman et al. 2003, 2006; Bergman 2007; Grossenbaugh et al. 2011). A recent study also found high expression of tyrosinase in equine melanomas. On the basis of this information, tyrosinase-targeted immunotherapies have been considered in horses (Phillips et al. 2012); however, their clinical application in the treatment of equine melanomas is still experimental.
Intra-lesional chemotherapy is often used alone or in combination with surgical excision or debulking to decrease the risk of recurrence. Intra-lesional application offers the advantage of decreasing the total administered dose of the chosen chemotherapeutic agent, thus minimising its adverse effects while preserving its efficacy (Théon 1998; Hewes and Sullins 2006). A variety of neoplastic masses in horses can be successfully treated with intra-lesional chemotherapy, consisting of cisplatin in oily emulsion (Théon et al. 1993) or in beads (Hewes and Sullins 2006), alone or in combination with surgical excision.
Intra-lesional chemotherapy with cisplatin is effective on equine extra-ocular dermal melanomas in 81% of the cases (Hewes and Sullins 2006; Théon et al. 2007). Carboplatin (cis-diammine [1,1-cyclobutanedicarboxylato] platinum [II]) is an analogue of cisplatin and it is a widely used chemotherapeutic agent with demonstrated activity against numerous solid tumours in man (Go and Adjei 1999).
Systemic carboplatin is effective in 11–19% human patients with advanced stage malignant melanoma (Evans et al. 1987; Casper and Bajorin 1990; Chang et al. 1993). Carboplatin is also effective in vitro in melanoma cells in dogs (Knapp et al. 1995). In a retrospective study of 27 cases of malignant melanoma in dogs, systemic carboplatin induced a response in 28% of the patients (Rassnick et al. 2001).
In the case reported here, intra-lesional chemotherapy was elected since the cleanness of the margins was questionable at the time of surgical excision and carboplatin was chosen due to immediate availability.
Swelling and inflammation have been described as potential complications of intra-lesional chemotherapy (Théon et al. 1994); however, they were not observed in the reported case.
Melanomas are relatively rare in the eye adnexa compared to other locations in horses (Giuliano 2010) and none have been reported on the third eyelid. Primary neoplasia of the nictitans is well recognised, with squamous cell carcinoma being the most common (Lavach and Severin 1977; Sundberg et al. 1977; Gearhart et al. 2007; Mathes et al. 2011). To our knowledge, this is the first report of a melanoma affecting the conjunctiva of the third eyelid in a horse.