Diagnostic challenges of metastatic spindle cell melanoma on fine-needle aspiration specimens

Authors


  • Presented at the 96th Annual Meeting of United States and Canadian Academy of Pathology, San Diego, California, March 24–30, 2007.

Abstract

BACKGROUND

Spindle cell melanoma is a morphologic variant of melanoma that can be difficult to diagnose on specimens obtained via fine-needle aspiration (FNA). Published cytology studies concerning this entity were based for the most part on small series. In the current study, a large series of metastatic spindle cell melanoma is described and the diagnostic pitfalls present in FNA samples addressed.

METHODS

The authors retrospectively reviewed the cytologic features of 81 metastatic spindle cell melanoma specimens obtained from 67 patients. Corresponding primary tumors or metastatic tumors taken elsewhere from the same patient were also evaluated.

RESULTS

The cytologic smears were mostly cellular and comprised of predominantly spindle tumor cells that frequently formed cohesive fascicles or whorls intermingled with scattered epithelioid tumor cells. The classic cytologic characteristics of conventional melanoma (predominantly dyshesive cellular distribution, cytoplasmic melanin pigments, intranuclear pseudoinclusions, macronucleoli, and binucleation or multinucleation) were noted infrequently or, if present, were more readily found in coexisting epithelioid cells. Remarkably, 9% of the cases failed to demonstrate any of the above classic characteristics. In addition, spindle cells demonstrated a wide range of cytologic atypia, from deceptively bland cells resembling reactive fibroblasts to those indistinguishable from pleomorphic high-grade sarcomatous neoplasms. When the morphologic features were compared with those of the primary tumor or metastatic melanoma taken elsewhere from the same patient, cell type discrepancy was found in 20% of the cases in that the previous counterparts demonstrated the epithelioid cell type. Spindle cells also tended to lose immunoexpression of melanoma markers.

CONCLUSIONS

Spindle cell melanoma infrequently demonstrates the diagnostic cytologic features and immunoreactivity of conventional melanoma. Varying degrees of cytologic atypia and possible cell type differences from the primary counterpart or metastatic melanoma occurring elsewhere are additional sources of diagnostic challenges, especially in the metastatic setting. Familiarity with cytologic features, combined with clinical and immunoperoxidase findings, is required to avoid misinterpretation. Cancer (Cancer Cytopathol) 2008. © 2008 American Cancer Society.

Melanoma is a common malignancy that occurs over a wide age range. It is also notorious for its strong tendency to metastasize to virtually any sites within the body and at any time after the primary tumor has been discovered. Some melanomas may take several decades to develop metastases,1 and others may present as a metastasis of unknown primary origin.2 Moreover, melanoma can display a wide range of cytologic appearances, thus mimicking a variety of other tumors.2, 3

The accurate diagnosis of metastatic melanoma has important prognostic and therapeutic implications. Fine-needle aspiration (FNA), a simple, fast, minimally invasive, and cost-effective procedure, is often used as an initial diagnostic tool in the evaluation of suspected metastatic or recurrent disease. The majority of the earlier descriptions of FNA findings of melanoma were based on features of epithelioid melanoma or on all types of melanoma as a whole.4–14 Characteristic cytologic features that have been reported to date are hypercellular aspirates, dispersed cellular distribution, epithelioid or plasmacytoid cells with eccentric nuclei, cytoplasmic melanin pigments, intranuclear pseudoinclusions, macronucleoli, and binucleation or multinucleation.4–14 Although the majority of melanomas are easily diagnosed, melanomas with unusual cytologic features can cause a diagnostic dilemma, particularly if the cells are sampled via FNA in a metastatic setting.

Spindle cell melanoma (including desmoplastic melanoma) is a morphologic variant of melanoma. Its reported incidence varies from 3% to 14%.4–6, 10–13, 15 However, to our knowledge, the FNA features of this variant have not been described in detail in the literature, and most have been reported as single case reports or small series.4, 5, 7, 12, 13, 16, 17 The misdiagnosis of a spindle cell melanoma is not uncommon, even on histopathologic examination.3 In the current study, we retrospectively reviewed the cytologic features and immunocytochemical findings of 81 FNA samples of metastatic spindle cell melanoma and discussed the potential diagnostic pitfalls in this setting.

MATERIALS AND METHODS

Using the pathology files of the University of Texas M. D. Anderson Cancer Center in Houston, we searched for FNA cases in which a diagnosis of spindle cell melanoma or melanoma with spindle cell features was rendered between January 2000 and July 2006. We retrospectively identified 81 cases of metastatic or recurrent disease from 67 patients for whom slides were available for review. In 10 patients, 2 or 3 synchronous metastatic lesions had been aspirated. Two patients underwent repeat aspirates on the same lesion. Clinical and/or radiologic information for all the patients was available to us for review. The study was conducted with the approval of the M. D. Anderson Cancer Center Institutional Review Committee.

Aspirates were obtained using a 22-gauge, 23-gauge, or 25-gauge biopsy needle. The aspiration was performed by a radiologist under imaging (computed tomography or ultrasonography) guidance when the lesion was deeply seated or by the reporting cytopathologist if the lesion was palpable. An average of 2 FNA passes was made for each case. Direct smears were air dried for Diff-Quik staining (Stat Lab, Lewisville, Tex) or fixed in modified Carnoy fixative (a 6:1 ratio of 70% ethanol to glacial acetic acid) for Papanicolaou (Pap) staining. The smears were assessed immediately by the cytopathologist for specimen adequacy. Cells obtained from the needle rinses were subjected to centrifugation, and the sediment was fixed in a 1:1 mixture of 95% ethanol to 10% formalin and embedded in paraffin to make cell blocks. The cell blocks were then sectioned and stained with hematoxylin and eosin.

Immunostaining was performed on cell block sections or on destained Pap-stained direct smears at the time of cytologic diagnosis in 36 of the 81 cases (44%): 10 on cell block sections and 26 on smears. We used the EnVision+ system and peroxidase detection methods in an autostainer (Dako Corporation, Carpinteria, Calif). The primary antibodies used were those against S-100 (1:40; BioGenex, San Ramon, Calif), HMB-45 (1:50; Dako Corporation), MART-1 (clone Ab-3 [1:100]; Neomarker/Labvision, Fremont, Calif), and Melan-A (clone A103 [1:50]; Neomarker/Labvison).

A final diagnosis of melanoma was rendered based on correlation of the cytologic features with the immunocytochemical findings and clinical and/or radiologic information. In addition, morphologic comparison of the metastatic tumors with their primary tumors or with metastatic tumors taken elsewhere from the same patient was attempted.

We retrospectively analyzed the cytologic features of each case for background content, cellularity, architecture of the cell arrangement, cell shape, and cytoplasmic and nuclear characteristics.

RESULTS

The patient population consisted of 44 men and 23 women. The mean age of the patients at the time of diagnosis of metastatic melanoma was 52 years (range, 17–81 years). The mean size of the tumor was 2.1 cm (range, 0.5–4.0 cm). The sites aspirated were soft tissue (35 patients), lymph node (22 patients), lung (12 patients), liver (5 patients), pancreas (4 patients), breast (1 patient), parotid gland (1patient), and bone (1 patient). The majority of metastases were discovered during clinical and radiologic surveillance. The mean interval between diagnosis of the primary melanoma and sampling of the metastatic tumors was 36.5 months (range, 1–217 months). All 67 patients in the current study had a known history of melanoma.

Although all cases demonstrated a predominance of spindle cells, in some cases variable amounts of epithelioid tumor cells were scattered among the spindle cells. Based on spindle cell composition, we grouped cases into spindle cell (SC) type when spindle cells constituted ≥90% of tumor cells and mixed cell (MC) type for the remaining cases. In the current study, 62 cases (77%) were SC type and 19 (23%) were MC type.

The cytologic features of the 81 cases are summarized in Table 1. We found that in most cases, the smears were moderately to highly cellular in both the SC and MC types. Compared with the MC type, the SC type tended to demonstrate more numerous and larger cohesive tissue fragments, which were comprised of interlacing fascicles or whorls of spindle cells (Fig. 1). The epithelioid cells, if identified, were mostly arranged singly or in a dispersed pattern. The cytologic features of the spindle cells were characterized by short or slender spindle cells with poorly defined cell borders, and variable amounts of cytoplasm with bipolar, thin, long cytoplasmic projections that were often curved or twisted at the ends. Rarely, the tumor cells had multiple projections, resulting in a dendritic appearance. Occasional bare nuclei were noted. Cytoplasmic melanin pigments of the spindle cells were variably observed as dusky and brown fine granules on Pap stain and blue-black on Diff-Quik stain. The nuclei of the spindle cells varied from oval to slender, and were occasionally wavy with a convoluted or folded nuclear membrane, which gave the appearance of nuclear grooving or angulation (Fig. 2). Intranuclear pseudoinclusions were variably noted, and were observed more frequently in coexisting epithelioid cells (Fig. 3A, inset). The majority of tumors demonstrated mild or moderate nuclear pleomorphism. We found that some tumor cells were extremely bland and uniform, resembling those of fibroblasts (Figs. 3 and 4), whereas others were markedly pleomorphic and bizarre, simulating pleomorphic, high-grade, sarcomatoid neoplasms (Fig. 5). Binucleated or multinucleated cells occasionally were found in the spindle cells, and these nuclei were usually placed in a row along the long axis of the cell (Fig. 5, inset). The chromatin was granular and delicate, and the nucleolus was mostly inconspicuous to visible, but occasionally was prominent.

Figure 1.

A smear of spindle melanoma cells arranged in a predominantly cohesive and fascicular pattern (Papanicolaou stain, ×200).

Figure 2.

A smear of spindle melanoma cells demonstrating longitudinal nuclear grooves and ill-defined cell borders admixed with scattered epithelioid cells (Papanicolaou stain, ×400). Inset: nuclear groove and cytoplasmic vacuoles (Diff-Quik stain, ×400).

Figure 3.

A smear of spindle cell melanoma demonstrating thin spindle cells with mild nuclear pleomorphism (A: Papanicolaou stain, ×200; B: H & E stain, ×200). Inset: intranuclear pseudoinclusions are more commonly observed in coexisting epithelioid cells (Papanicolaou stain, ×200).

Figure 4.

A smear of spindle melanoma cells in a lymph node demonstrating extremely bland nuclear features with evenly distributed granular chromatin, resembling fibroblasts. The poorly defined, pale cytoplasm has bipolar, thin, and long projections (A: Papanicolaou stain, ×100; B: Papanicolaou stain, ×200).

Figure 5.

A smear of spindle cell melanoma demonstrating marked nuclear atypia and pleomorphism, resembling pleomorphic high-grade sarcomatoid neoplasm (Papanicolaou stain, ×400). Inset: a binucleated spindle tumor cell in which nuclei were placed in a row along the long axis (Papanicolaou stain, ×400).

Table 1. Cytologic Features of 81 Cases of Melanoma With Spindle Cells*
FeatureSpindle cell type (n = 62)Mixed cell type (n = 19)Total (n = 81)
  • *

    Data are shown as the number (percentage).

Cellularity
 Moderate to high48 (77)15 (79)63 (78)
 Low14 (23)4 (21)18 (22)
Architecture
 Predominantly dyshesive9 (15)10 (53)19 (24)
 Mixed dyshesive and cohesive35 (56)7 (37)42 (52)
 Predominantly cohesive18 (29)2 (11)20 (25)
Cytoplasmic melanin pigment7 (11)5 (26)12 (15)
Nuclear pleomorphism
 Mild25 (40)4 (21)29 (36)
 Moderate23 (37)8 (42)31 (38)
 Marked14 (23)7 (37)21 (26)
Intranuclear pseudoinclusion12 (19)6 (32)18 (22)
Binucleation/multinucleation7 (11)11 (58)18 (22)
Macronucleolus17 (27)11 (58)28 (35)

Overall, the characteristic features of conventional melanoma (ie, a predominantly dyshesive cellular arrangement, cytoplasmic melanin pigments, intranuclear pseudoinclusions, macronucleoli, and binucleation or multinucleation) were more subtle and less frequently found in cases of the SC type compared with MC type cases (Table 1). If present, these features tended to be associated with coexisting epithelioid cells. It is interesting to note that 7 of these 81 cases (9%) did not exhibit any of these characteristics.

Occasional mitotic figures were found in 12 cases (15%) and small foci of necrosis were noted in 10 cases (12%). Cytoplasmic vacuoles were more obvious in slides stained with Diff-Quik than in Pap-stained slides. The vacuoles were found in 17 of the 57 cases for which Diff-Quik-stained slides were available (30%), and tended to be fewer and smaller in the spindle cells. These features were identified with similar frequency in cases of both SC and MC types.

Morphologic comparison of cell type between the current study cases and previously obtained specimens of primary tumor or metastatic melanoma taken elsewhere from the same patient was performed in 49 cases. We found that 39 of the 49 cases (80%) demonstrated cell types that were comparable to their previous counterparts, and 10 cases (20%) exhibited different cell types in that the previous counterparts were epithelioid in shape (Table 2). Such morphologic comparisons had facilitated the diagnosis of melanoma in 25 of the current study cases (31%) and all these cases displayed cellular features comparable to those of the previous counterparts.

Table 2. Comparison of Cell Type Between Current Cases and Previous Counterparts From the Same Patients
Previous cell typeCurrent cell type
Spindle cell (n = 62)Mixed cell (n = 19)
Spindle cell225
Mixed cell93
Epithelioid55
Not specified266

At the time of cytologic diagnosis, immunoperoxidase studies were performed in 36 of the 81 cases (44%). Positive staining for S-100 was found in 67% (Fig. 6), whereas 50% of cases were positive for HMB-45, 31% were positive for Melan-A, and 18% were positive for MART-1 (Table 3). Seventy-two percent of tumors stained positively for ≥1 of these 4 markers.

Figure 6.

(A) An example of S-100 staining on a previously Papanicolaou-stained smear of metastatic spindle cell melanoma demonstrating weak S-100 staining in rare tumor cells (S-100 immunostaining, ×100). (B) An example of S-100 staining on cell block material from metastatic spindle cell melanoma demonstrating readily identified positive tumor cells (S-100 immunostaining, ×200).

Table 3. Immunostaining Results of Melanoma Markers in 36 Examined Cases*
ImmunomarkerSpindle cell type (n = 28)Mixed cell type (n = 8)Total (n = 36)
  • *

    Data are shown as the number of positive stains (percentage).

Positive for ≥1 of the following 4 markers20/28 (71)6/8 (75)26/36 (72)
S-10015/21 (71)3/6 (50)18/27 (67)
HMB-453/8 (38)2/4 (50)6/12 (50)
Melan-A4/11 (36)0/2 (0)4/13 (31)
MART-11/9 (11)1/2 (50)2/11 (18)

DISCUSSION

FNA is often used to document recurrent or metastatic disease. A diagnosis of melanoma can be readily made when the characteristic cytologic features of conventional melanoma are present: hypercellular samples, variably pleomorphic epithelioid/plasmacytoid cells arranged in a predominantly dyshesive pattern, cytoplasmic melanin pigments, intranuclear pseudoinclusions, macronucleoli, and binucleated or multinucleated tumor cells. Spindle cell melanoma (including desmoplastic melanoma) is a variant that often leads to misdiagnosis but to our knowledge has not been well described in the cytopathology literature published. In the current study, we found that spindle melanoma cells demonstrated varying degrees of nuclear atypia and infrequently displayed the above-mentioned characteristic features. Moreover, these features, when present, were detected more readily in coexisting epithelioid tumor cells than in spindle cells.

The presence of melanin pigments is a valuable indicator with which to justify the diagnosis of melanoma. The incidence of cytoplasmic pigmentation in FNA samples varies in different studies, ranging from 30% to 80%.4, 5, 7, 8, 10, 12–14 In the current study, only 15% of the cases had recognizable cytoplasmic melanin. When melanin pigments were identified, they were often quite subtle and sparse and thus easily overlooked. Intranuclear pseudoinclusions, macronucleoli, and binucleation or multinucleation, although not specific, can provide useful clues to the tumor origin. The frequency of each feature in melanoma was reported to be approximately 50%.5, 7, 10, 12, 13 We found that these features were noted less frequently in the spindle cell melanomas (22%, 35%, and 22%, respectively), and 7 of the current study cases (9%) failed to exhibit any of the characteristic features. In addition, the cells of spindle cell melanoma tended to form cohesive microtissue fragments with a fascicular or storiform arrangement. Such features can be the source of diagnostic pitfalls.

Previous studies have documented findings similar to those noted herein. Woyke et al.12 and Murali et al.14 reported that spindle cell melanoma tended to form aggregated cell groups or tissue fragments. Perry et al.5 reported that pleomorphism was more frequent in epithelioid melanoma, whereas bland nuclei and inconspicuous nucleoli were far more frequent in spindle cell melanoma. Similarly, Nasiell et al.13 found that cytoplasmic melanin pigmentation, vacuolization, nucleoli, and intranuclear pseudoinclusions were observed less frequently in spindle cell melanoma compared with conventional melanoma.

FNA specimens of metastatic spindle cell melanomas aspirated from various anatomic sites could be mistaken for metastatic lesions from other neoplasms or reactive lesions with similar features, especially when a reliable clinical history is unavailable and the tumor cells are predominantly or “purely” spindle shaped without the characteristic features of melanoma. We observed a wide range of pleomorphism and cytologic atypia within the spindle cells in the current study. Approximately 30% of the current study cases demonstrated generally mild nuclear pleomorphism. Some tumors demonstrated extremely bland spindle cells with uniform, slender, and wavy nuclei, reminiscent of fibroblasts in a reactive process or benign spindle cell tumors (eg, schwannoma, neurofibroma, dermatofibroma, benign fibrohistiocytoma, and fibromatosis). It is conceivable that a recurrent melanoma with such morphologic characteristics found adjacent to a recent excision site could be mistaken for an exuberant mesenchymal repair with fibroblastic proliferation or scar tissue. Chhieng et al.17 reported a case in which bland spindle melanoma cells metastasized to a lymph node and the FNA sample was initially misdiagnosed as a reactive lymph node with fibrosis. Bland spindle tumor cells with grooved nuclei and inconspicuous nucleoli may bear some resemblance to centrocytes of the lymph node.3 Conversely, spindle cell melanoma can also demonstrate significant nuclear atypia and pleomorphism, simulating pleomorphic high-grade sarcoma or sarcomatoid carcinoma. Erroneous diagnoses have been reported because the morphologic features of spindle cell melanoma overlap with those of malignant fibrohistiocytoma, fibrosarcoma, malignant peripheral nerve sheath tumor, leiomyosarcoma, and synovial sarcoma.3 It is noteworthy that, although the presence of cytoplasmic pigmentation is an important clue in establishing a melanocytic nature, the pigments can occasionally be identified in melanotic schwannoma and melanotic malignant peripheral nerve sheath tumor.18, 19 Moreover, a peripheral nerve sheath tumor can express S-100, and it occasionally demonstrates melanosomes under an electron microscope, thus making a differential diagnosis even more challenging.3, 14

Misinterpretation usually occurs when the diagnosis is based solely on cytomorphologic features and an appropriate clinical history and ancillary workup are not available. It is important to note that some metastatic melanomas may present with an unknown primary origin because of spontaneous regression or misdiagnosis of the primary melanoma, and others may occur decades after surgical treatment of the primary melanoma.1, 2 In the current series, the longest interval between the excision of the primary melanoma and the development of metastasis was 217 months (18 years). FNA diagnosis of spindle cell melanoma in these patients could be challenging if the classic morphologic features of melanoma are lacking because a remote history may lead to a favorable consideration of a second primary tumor. Furthermore, patients with melanoma have an increased risk of developing a second, unrelated primary malignancy.20, 21 Therefore, one needs to consider the possibility of a new primary malignancy, even when a history of melanoma is known.

Familiarity with the cytologic features of spindle cell melanoma is very important in rendering an accurate diagnosis. Knowledge of the morphologic features of the previous counterpart (either the primary tumor or metastatic melanoma occurring elsewhere) will further increase the diagnostic certainty. In the current series, the diagnosis of 31% of cases was facilitated by a morphologic comparison with previous pathologic materials. However, it is interesting to note that a discrepancy in cell type may occur. We found that 20% of metastatic spindle cell melanomas (SC type or MC type) had epithelioid morphology in their previous counterparts. Such a discrepancy has been described previously,2, 3, 22 and can cause a diagnostic dilemma. Although it might be prudent to consider the possibility of a new primary tumor, an immunoperoxidase study usually clarifies the diagnosis.

The markers S-100, HMB-45, Melan-A, and MART-1 are commonly used to confirm a melanocytic nature.2, 3, 7, 13, 23–26 S-100 is a sensitive marker that labels primary and metastatic melanomas, including the spindle cell variant.7, 13, 24 However, S-100 expression can also be detected in a variety of mesenchymal tumors (especially in peripheral nerve sheath tumors) and occasionally in carcinomas.13, 27–30 Conversely, false-negative S-100 staining may be encountered if the FNA smears used for staining are fixed in an alcohol-based solution.31 In the current study, we also noted that, compared with cell block material, Pap-stained smears demonstrated positive S-100 staining less frequently. If positive cells were found in smears, they often demonstrated weak staining in rare cells (Fig. 6). HMB-45, Melan-A, and MART-1 are relatively specific but less sensitive markers for melanomas. Spindle melanoma cells frequently lose expression of these markers except in foci of epithelioid cells.3, 32 Therefore, using a panel of antibodies may improve the certainty of distinguishing between a spindle cell melanoma and its mimickers. In the current study, positive staining for S-100, HMB-45, Melan-A, and MART-1 was found in 67%, 50%, 31%, and 18%, respectively, of the cases examined. Seventy-two percent of the tumors demonstrated at least 1 of these 4 markers. It is possible that some of the current study cases may have demonstrated false-negative immunoperoxidase staining as a result of insufficient FNA sampling, as well as the destaining procedure and alcohol-based fixative used in previously Pap-stained smears. Cytokeratin is a commonly used discriminate marker for melanoma. However, aberrant expression of cytokeratin has been described in up to 10% of melanomas, and it is more commonly observed in metastatic tumors.3, 13, 30 If it is noted, a diagnosis of sarcomatoid carcinoma needs to be excluded. Another aberrantly expressed marker that is found occasionally in spindle cell melanoma is smooth muscle actin,33 which can lead to the misinterpretation of spindle cell melanoma as a tumor of smooth muscle origin. Lastly, electron microscopy may be of diagnostic help if premelanosomes or melenosomes are identified, but these are often sparse and atypical in spindle cell melanoma.

In summary, spindle cell melanoma may mimic other spindle cell lesions because the characteristic features of conventional melanoma may be lacking and there may be a variable degree of cytologic atypia. Other potential pitfalls are the variable loss of expression of melanocytic markers and an occasional discrepancy in cell type between metastatic spindle cell melanomas and their previous counterparts. Knowledge of a prior history of melanoma is helpful to avoid an erroneous interpretation. In addition, a full awareness of the FNA features of spindle cell melanoma and the findings of a panel of immunomarkers greatly improve the diagnostic certainty of metastatic spindle cell melanoma by FNA.

Ancillary