Signalment:  

12-year-old intact male Pug, dog (Canis familiars).The dog had a recent history of an oral melanoma, edema (not further specified), and cavitary effusion (not further specified). The dog had multiple biopsies of the oral mass that was on the left mandible, which was confirmed to be malignant melanoma. The mass with incomplete margins was removed along with teeth 304-308. The dog was ultimately euthanized.

Gross Description:  

The entire body of the dog was submitted in a state of fair to good postmortem preservation. The dog was in a good body condition with a moderate amount of subcutaneous and intraabdominal adipose tissue. There were no visible teeth in the 300 arcade (lower left) within the mandible. There was a locally invasive 5.5 cm x 4 cm x 5.5 cm firm, tan to white, multilobular mass encompassing the left side of the oral cavity involving both the mandible and maxilla. The mass surrounding the mandible extended caudally from the rostral aspect of the body of the mandible to the ramus, slightly across the midline in the caudal intermandibular region, and dorsally surrounding the caudal most aspect of the maxilla including the remaining maxillary molar (presumably tooth 210). The mass completely surrounded the mandible; however, it was not attached to the mandible. The left mandible was markedly thinned with a complete, mid-body, transverse fracture present. On cut section, the mass was tan to white and semifirm with a tan, soft, gelatinous central region and occasional cavities that oozed a small amount of yellow to green, semi-viscous, opaque material. 

The lungs were diffusely mottled, tan to dark red and crepitant. There were multifocal, round, tan, slightly raised nodules ranging from pinpoint to 4 mm diameter throughout all lung lobes, but affecting less than 1% of the pulmonary parenchyma. One pedunculated tan nodule was present on the right caudal lobe. On cut section the pulmonary nodules were tan.

Histopathologic Description:

Compressing the adjacent salivary gland and submucosa as well as infiltrating and dissecting between the adjacent skeletal muscle and collagen bundles, and occasionally infiltrating the basal epithelial layer (junctional activity variable by section), there is a densely cellular, multilobular, poorly demarcated, unencapsulated, infiltrative neoplasm composed of two populations of cells. One population of cells is arranged in cords, trabeculae, and packets and supported by a fine fibrovascular stroma. The neoplastic cells are polygonal to columnar with variably distinct cell borders, a moderate amount of eosinophilic cytoplasm, and one round to oval nucleus with finely stippled chromatin and one to three prominent nucleoli. The second population of cells is arranged in tightly arranged packets and supported by a fine fibrovascular stroma. The neoplastic cells are polygonal with indistinct cell borders, a large amount of clear to lightly eosinophilic, vacuolated cytoplasm, and one round to oval nucleus with finely stippled chromatin and one to two variably prominent nucleoli. Overall the neoplastic cells exhibit a moderate degree of anisocytosis and anisokaryosis with a mitotic rate of approximately 35 in 10 high power fields (400x). Multifocally within the neoplasm, the stroma is brightly eosinophilic and hyalinized. Multifocally, there are variably sized regions of coagulative to lytic necrosis characterized by eosinophilic and basophilic karyorrhectic cellular debris and a pale eosinophilic background with faded cellular outlines and nuclear details. Along the periphery of the neoplasm and admixed within the regions of necrosis, there are moderate numbers of degenerate and non-degenerate neutrophils, nuclear streaming, a small amount of deeply basophilic granular material (mineral), and small numbers of mixed bacterial colonies. The bacterial colonies are characterized as basophilic cocci and eosinophilic short rods. Scattered throughout the neoplasm are moderate numbers of individual apoptotic/necrotic cells with pyknotic nuclei and karyorrhectic debris. Throughout the neoplasm, numerous small and medium caliber blood vessels are expanded by moderate numbers of erythrocytes (congestion). Multifocally throughout the neoplasm, there are small to moderate aggregates of extravasated erythrocytes (hemorrhage). Along the periphery of the neoplasm and extending into the surrounding collagen bundles, there are small aggregates of lymphocytes and plasma cells. Within the adjacent muscle, there is myofiber degeneration and necrosis characterized by myofiber loss, variation in myofiber size, and loss of cross-striations. Within these same regions along the periphery of the neoplasm, there is moderate collagenolysis characterized by a loss of organization, collagen bundle fragmentation, loss of eosinophilia and increased basophilia. There are multifocal areas of pigmentary incontinence. Multifocally there is a loss of the overlying oral epithelium (ulceration). 

Immunohistochemistry for Melan-A, S100, chromogranin A, and synaptophysin were prepared at the University of Minnesota Veterinary Diagnostic Laboratory. Less than 2% of neoplastic cells within the periphery of each lobule had strong intracytoplasmic immunopositivity and less than 10% of the neoplastic cells throughout the mass had weak to moderate intracytoplasmic immunopositivity for Melan-A. Approximately 50% of the neoplastic cells had weak intracytoplasmic immunopositivity for Chromogranin A. The neoplastic cells were immunonegative for S100 and Synaptophysin. 

Morphologic Diagnosis:  

Oral cavity, amelanotic melanoma with neuroendocrine differentiation, focally extensive (with metastasis to lungs and left mandibular pathologic fracture).

Lab Results:  

There were no ancillary tests performed.

Condition:  

Amelanotic melanoma

Contributor Comment:  

The two main types of melanocytic tumors are melanocytoma and malignant melanoma.(8) These tumors are derived from melanocytes that originate as melanoblasts from neural crest ectoderm.(8) Melanocytomas are the benign neoplasm of melanocytes and are most often heavily pigmented.(8) As the name suggests, malignant melanoma is the malignant tumor of melanocytic origin and is often used synonymously with melanoma. Melanocytic tumors can be found in multiple locations; however, the integument (cutaneous) and oral cavity are common sites. Cutaneous tumors in dogs are often benign. The cutaneous form of melanoma is commonly seen in gray horses and swine.(3)

In dogs, malignant melanomas are the most common oral tumor and have variable breed and gender predilection.(3,7-9) These tumors are often heavily pigmented and grossly appear black; however, in some cases, these tumors are variably pigmented or completely unpigmented (amelanotic), and appear grossly white.(3,7,8) The degree of pigmentation does not indicate biologic behavior or aid in prognosis.(3,8) Malignant melanomas often metastasize, with the regional lymph nodes being the most common site, but also commonly spread to distant sites (ex. lung) through the blood and lymphatics.(3,9) Histologically, the cell morphology of malignant melanomas can range from round/polyhedral/epithelioid cells to spindloid cells to mixed (a combination of the two)(3,8); however, one defining histologic feature of the these tumors is often junctional activity(4), which can be seen as tumor infiltration crossing between the basal epithelium of the mucosa and the submucosa. 

Most often, well-differentiated round/polygonal/epithelioid cell melanocytic tumors with abundant pigmentation are easily diagnosed; however, amelanotic and spindloid appearing tumors are quite often diagnostically challenging. Special stains [Fontana-Masson, 3,4-dihydroxyphenylalanine (DOPA)], immunohistochemistry, and/or electron microscopy can be used to aid in the diagnosis of amelanotic tumors.(1,3,5,7,8,10) Amelanotic melanomas are generally immunopositive for tryrosinase-related proteins 1 and 2 (TRP-1 and TRP-2), Melan-A, melanocytic antigen PNL2, HMB-45, microphthalmia transcription factor (MiTF), S100, tyrosine hydroxylase, tyrosinase, and vimentin(3,5,7-10); however, according to Smedley et al. PNL2, Melan-A, TRP-1 and TRP-2 as a group provide the most diagnostic information for canine oral amelanotic melanomas. 

In human medicine, melanocytic tumors are often further classified allowing for integration of morphology, location, and biologic behavior; however, this classification system is not currently applied in veterinary medicine.(8) According to Eyden et al., while not currently a subclassification of malignant melanoma, neuroendocrine differentiation is a rarely seen variant. The three currently reported human cases of this variant had predominantly epithelioid cell morphology, variable pigmentation, and immunopositivity either in the primary tumor or metastases for both melanocytic and neuroendocrine markers. Ultrastructurally, the assessed cases (2 of 3) revealed neuroendocrine granules and no melanosomes.(6) Although electron microscopy was not performed on the current case, the cell morphology, architecture, and immunophenotype are consistent with an amelanotic melanoma with neuroendocrine differentiation; which has currently not been reported in dogs.

JPC Diagnosis:  

Fibrovascular tissue adjacent to salivary gland: Malignant amelanotic melanoma.

Conference Comment:  

Melanomas are often referred to as the great imitator because of their ability to display a variety of cytologic features thanks to their common embryologic roots with both neural and epithelial tissues. This case nicely illustrates the point, due to variation is cellular appearance across various regions of the neoplasm. In some sections, there is a portion of the neoplasm which exhibits junctional activity, a feature which lends additional credence to the morphologic diagnosis on HE section. The list of other neoplasms that share this characteristic with melanomas is short, usually limited to histiocytomas and epitheliotropic (T-cell) lymphomas. 

Nuclear atypia and mitotic activity is directly correlated with prognostic behavior in melanomas at all locations. Immunohistochemistry has recently been evaluated as an additional prognostic tool. Ki67, a protein expressed only in growth and mitotic phases of the cell cycle, is considered a measure of tumor growth fraction and its expression has been correlated with outcome in canine melanomas comparable to nuclear atypia and mitotic index.(2) Additionally, immunodetection of RACK1, a protein expressed at high levels in melanocytes, may be a specific marker aiding in not only identifying melanomas but also offering progonostic value as its expression is also consistently correlated with nuclear atypia and mitotic index.(4)

References:

1 Banerjee SS, Harris M. Morphological and immunophenotypic variations in malignant melanoma. Histopathology. 2000;36:387-402.

2. Bergin IL, Smedley RC, Esplin DG, Spangler WL, Kiupel M. Prognostic evaluation of Ki67 threshold value in canine oral melanoma. Vet Pathol. 2011;48(1):41-53.

3. Brown CC, Baker DC, Barker IK. Alimentary system. In: Maxie MG, ed. Jubb, Kennedy, and Palmer's Pathology of Domestic Animals. 5th ed. Toronto: Saunders/Elsevier; 2007:29-30.

4. Campagne C, Jule S, Alleaume C, et. al. Canine melanoma diagnosis: RACK1 as a potential biologic marker. Vet Pathol. 2013;50(6):1083-1090.

5. Choi C, Kusewitt DF. Comparison of Tyrosinase-related Protein-2, S-100, and Melan A Immunoreactivity in Canine Amelanotic Melanomas. Vet Pathol. 2003;40:713-718.

6. Eyden B, Pandit D, Banerjee SS. Malignant melanoma with neuroendocrine differentiation: clinical, histological, immunohistochemical and ultrastructural features of three cases. Histopathology. 2005;47:402-409.

7. Gelberg HB. Alimentary system and the peritoneum, omentum, mesentery, and peritoneal cavity. In: Zachary JF, McGavin MD, eds. Pathologic Basis of Veterinary Disease. 5th ed. St. Louis, MO: Elsevier; 2012:329.

8. Head KW, Cullen JM, Dubielzig RR, et al. Histological Classification of Tumors of the Alimentary System of Domestic Animals. Schulman FY, ed. 2nd series, Vol.10. Washington, DC: AFIP, CL Davis foundation, and WHO; 2003:33-35.

9. Ramos-Vara JA, Beissenherz ME, Miller MA, et al. Retrospective study of 338 canine oral melanomas with clinical, histologic, and immunohistochemical review of 129 cases. Vet Pathol. 2000;37:597-608. 

10. Smedley RC, Lamoureux J, Sledge DG, Kiupel M. Immunohistochemical Diagnosis of Canine Oral Amelanotic Melanocytic Neoplasms. Vet Pathol. 2011;48(1):32-40.

Click the slide to view.

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3-1. Head

3-2. Soft tissue, jaw

3-3. Soft tissue, jaw

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