TEACHING CASE 3

Relevant clinical history:
The patient is a 56 year old male with no significant previous medical history who presented with acute hemorrhage into the right frontal lobe. The blood clot was evacuated surgically, and approximately 4 ml of organized, dark burgundy material was submitted to the pathology lab.

Histologic examination of the material revealed that it was largely a cellular, although there were, in fact, fragments of tissue that looked like cerebral cortical tissue containing clusters of malignant cells. The differential diagnosis of this malignancy is essentially that of the 'large cell undifferentiated' malignant neoplasm where the differential diagnosis includes carcinoma, lymphoma, melanoma, and (round cell) sarcoma. In the case of a lesion in the brain, such as this, the diagnosis of malignant glioma should also be entertained. A simple panel of antibodies can be helpful in distinguishing among these diagnostic possibilities.

The questions posed by the referring pathologist are: (a) can this tumor be more precisely subclassified?; and (b) where is its primary site? Immunohistochemical studies can address both of these questions.

CD43	Antibodies to CD45 and CD43 (shown in the figure) were both negative. Antibodies to CD43 were included because one important diagnosis to consider in the differential of 'large cell undifferentiated' malignant neoplasms is anaplastic large cell lymphoma (ALCL), and a significant subset of the latter can be CD45-negative. CD43 is an excellent and highly sensitive marker of ALCL, and also has the advantage of marking most leukemic infiltrates as well. Note the CD43-positive lymphocytes and macrophages in the image; the tumor cells, however, are negative.
GFAP	Antibodies to GFAP, the intermediate filament protein of glial cells and tumors, are negative on the cells of interest. Note, however, the considerable 'background' which is actually immunostaining of the glial cells and processes in the cerebral cortical tissue. The tumor cells are GFAP-negative.
CYTOKERATIN	Antibodies to cytokeratin, the intermediate filament protein of epithelial cells and tumors, are negative on the cells of interest.

Having ruled out carcinoma, lymphoma and glioma, attention now turns to the possibility of the diagnosis of melanoma. HMB45, an antibody developed by Dr. Gown with a colleague, Dr. Vogel, in the mid-1980s, is still considered by many the 'gold standard' for the diagnosis of melanoma, although in our experience the diagnosis of melanoma can be assisted by incorporating a 'dream team' of several different anti-melanoma antibodies, including HMB45 as well as antibodies to tyrosinase and the MART1 gene product. The following images show that, using a cocktail of three monoclonal antibodies, the tumor cells are positively identified as representing metastatic melanoma.

MELANOMA COCKTAIL

Here are more detailed explanations of the three melanoma markers incorporated in our antibody cocktail:

• HMB45: gp100 is the name of the premelanosomal antigen identified by antibody HMB45 (as well as its 'sister' antibody, HMB50). Expression of this antigen is best considered a marker of the presence of these organelles rather than a particular cell type (e.g., melanocytic), because there are non-melanocytic, non-nevocytic processes that can contain premelanosomes and hence manifest HMB45/50-positivity. The latter include angiomyolipoma (as well as recently described malignant processes arising from angiomyolipomas) and lymphangioleiomyomatosis.
• Tyrosinase is a newly described melanoma-restricted marker that can help identify metastatic melanomas, including cells found in sentinal lymph nodes. (de Vries, T. J. et al. Cancer Res 57:3223-9, 1997. Hofbauer, G. F. et al. J Cutan Pathol 25:205-9, 1998.)
• The MART1 antigen is a melanocytic/nevocytic antigen different from the HMB-45-defined gp100 antigen. While comparable in sensitivity to HMB-45 for the detection of melanomas, unlike HMB-45, antibodies to MART1 identify most benign nevocytic/melanocytic tumors, particularly those within the dermis (de Vries TJ et al., Cancer Res 57:3223-9, 1997). Furthermore, the MART1 antigen, is expressed in adrenal cortical tumors (but not renal cell carcinoma). (Busam KJ et al. Am J Surg Pathol 22:57-63, 1998).

FINAL DIAGNOSIS: METASTATIC MALIGNANT MELANOMA