- Email: [email protected]
Melanocytic lesions – Staying out of trouble
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ACCEPTED MANUSCRIPT Melanocytic Lesions – Staying Out of Trouble
Thomas Brenn, MD, PhD, FRCPatha, b
of Pathology & Laboratory Medicine and Medicine and
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aDepartments
The Arnie Charbonneau Cancer Institute Cumming School of Medicine University of Calgary Diagnostic and Scientific Centre
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9 – 3535 Research Rd NW
Canada
of Pathology
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bDepartment
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Email: [email protected]
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University of Edinburgh
Western General Hospital
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Crewe Rd
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Calgary, AB, T2L 2K8
Edinburgh, EH4 2XU United Kingdom
Declarations of interest: none
ACCEPTED MANUSCRIPT Abstract The diagnosis of melanocytic tumors is notoriously difficult and represents one of the most challenging areas in surgical pathology associated with significant risk for litigation. One reason is the wide morphologic spectrum of melanocytic tumors and the fact that many
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histological features are shared by both benign melanocytic nevi and melanoma. Awareness of the many morphologic variations and variants of nevi and melanoma, their clinical setting, immunohistochemical phenotype and genetic profile is necessary for the correct diagnosis. The article discusses the features of three variants of melanoma, desmoplastic, acral lentiginous and vulvar mucosal lentiginous melanoma, and their benign melanocytic
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mimics with emphasis on distinguishing features.
Keywords
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Melanoma, Nevi, Melanocytic, Immunohistochemistry, Genetics, Diagnosis
ACCEPTED MANUSCRIPT Introduction The correct diagnosis of melanocytic lesions is one of the biggest challenges in dermatopathology and one of the most frequent causes for litigation against Pathologists. There are multiple explanations for this. Some relate to the accuracy or lack of important
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clinical data such as age, gender, anatomical location, clinical presentation and level of clinical suspicion. Others concern the type of biopsy. There is a trend to smaller and more superficial biopsies, which may not be representative of the entire lesion. Finally, the morphologic spectrum of melanocytic tumors is wide with overlapping features of melanoma and benign nevi creating significant diagnostic pitfalls. The following text
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contains a discussion of some of the variants of benign melanocytic nevi which may be
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difficult to separate from invasive melanoma. Although some of these nevi are rare their
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recognition is important to avoid overdiagnosis as melanoma.
1. Melanocytic tumors with stromal desmoplasia
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1.1 Desmoplastic melanoma
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Desmoplastic melanoma is an unusual but distinctive variant of invasive melanoma linked to chronic UV exposure. It shows specific clinical, histological, immunohistochemical and
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genetic features and behavior. Desmoplastic melanoma is a frequent cause for misdiagnosis, especially on partial samples, with a broad differential diagnosis. Clinically, desmoplastic melanoma shows a strong predilection for chronically sun-damaged skin of the head and neck, particularly the scalp, of elderly males. It presents as slowly growing flesh colored nodules and plaques. Clinically visible pigment is seen in a minority of tumors (1).
ACCEPTED MANUSCRIPT Due to its infiltrative growth pattern and neurotropism desmoplastic melanoma shows high local recurrence rates. The overall outcome of ‘pure desmoplastic melanoma’ appears to be favorable compared to conventional melanoma when adjusted for tumor thickness. The tumors also show reduced rates of metastasis to locoregional lymph nodes (2). In contrast,
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the outcome and lymph node metastatic rates of ‘mixed desmoplastic melanoma’ are similar to conventional melanoma.
Histologically, the tumors are poorly circumscribed and asymmetrical with a plaque-like growth pattern in the dermis and a diffusely infiltrative growth into underlying subcutaneous adipose tissue, skeletal muscle or fascia. On low-power inspection the tumors
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are broad, ill-defined and asymmetric (Fig 1a). So-called ‘pure desmoplastic melanoma’ is
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composed of spindle cells arranged singly, in small nests or short fascicles in a desmoplastic stroma (Fig 1b and c). The tumors are paucicellular although the overall cellularity is
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variable. Cytological atypia is usually mild with nuclear hyperchromasia, and the mitotic activity is low (Fig 1d). Peripheral lymphoid aggregates are present and are a good low-
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power diagnostic clue (Fig 1a). Perineurial infiltration is frequently identified. There may be
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an atypical lentiginous melanocytic proliferation involving the overlying epidermis, in some instances amounting to lentigo maligna. Classic melanoma in situ is only rarely encountered.
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A subset of tumors, so-called ‘mixed desmoplastic melanoma’, is characterized by an additional second, more conventional epithelioid or spindled invasive dermal component, characterized by a cellular and sheet-like growth and pronounced cytologic atypia (3, 4). By immunohistochemistry, ‘pure desmoplastic melanoma’ expresses S100, SOX10 and P75 consistently but they are typically negative for melan A, Mart1 and HMB45 (Fig 1e). P16 is expressed in a subset of desmoplastic melanomas (5).
ACCEPTED MANUSCRIPT Genetically, desmoplastic melanomas show frequent mutations in the NF-1 gene but mutations in BRAF, NRAS, GNAQ, GNA11 or KIT are absent (6, 7). The differential diagnosis of desmoplastic melanoma is wide and includes melanocytic and non-melanocytic tumors. Non-melanocytic entities, such as spindle cell squamous cell
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carcinoma, atypical fibroxanthoma, pleomorphic dermal sarcoma, fibrous histiocytoma, dermatofibrosarcoma protubertans and scars can readily be separated by the absence of S100 or SOX10 expression. In contrast the distinction from neurofibroma, especially plaquetype neurofibroma, is more challenging as both tumors share S100 and SOX10 expression. Lack of cytological atypia, absence of the desmoplastic stroma and strong and diffuse CD34
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expression are helpful morphologic and immunohistochemical clues. Furthermore, diffuse
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neurofibroma occurs in young adulthood and does not favor chronically sun-damaged skin. A number of benign melanocytic nevi that may be readily mistaken for desmoplastic
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melanoma are discussed in more detail.
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1.2. Melanocytic nevi with perineurial differentiation (perineuriomatous nevi)
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Melanocytic nevi with perineurial differentiation are a rare but histologically distinctive and potentially under-recognized variant of acquired nevi showing extensive neurotization with
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perineurial differentiation. They are easily mistaken for desmoplastic melanoma on histology (8, 9).
Clinically, they present in adulthood with a male predominance and wide anatomical distribution affecting the trunk, the extremities, the neck and scalp. Often there is a history of a longstanding pigmented lesion with recent change. Their behavior is entirely benign (8, 9).
ACCEPTED MANUSCRIPT Histologically, the tumors are relatively circumscribed but asymmetric (Fig. 2a). They involve the entire dermis and may extend into the superficial subcutaneous adipose tissue. Their growth pattern is biphasic with a common acquired compound or dermal nevus component in the superficial aspect (Fig. 2b). In the deeper reaches, the tumor is composed of uniform
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and bland spindle cells arranged in intersecting fascicles in a variably myxoid and desmoplastic stroma (Fig. 2c). In areas, the spindle cells show slender, wavy nuclei and long, bipolar cytoplasmic processes reminiscent of perineurial cells (Fig. 2d). There is no cytologic atypia or increased mitotic activity. Additional characteristic features include intratumoral hypocellular sclerotic nodules and peripheral lymphoid aggregates (2e and f).
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By immunohistochemistry, the conventional nevic component expresses S100 and melan A
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strongly and diffusely, while HMB45 expression is confined to the junctional and superficial dermal aspect only. The deeper spindle cell component is comprised of and admixture of
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S100-positive, melan A-negative cells and S100-, melan A-negative cells showing expression of the perineurial markers EMA, glut1 and claudin1(8) (Fig. 2g and h).
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Separation from desmoplastic melanoma is challenging on histology. Helpful features
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include the overall circumscription, the lack of cytological atypia, the presence of the distinctive hypocellular nodules and the presence of a common acquired compound or
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dermal nevus in the superficial aspects of the tumor. The clinical presentation further aids the distinction. The tumors have a longstanding history and they show no predilection for chronically sun-damaged skin of the elderly.
1.3. Desmoplastic melanocytic nevi A desmoplastic stroma may be seen in different subtypes of nevi including blue (sclerosing blue) and Spitz (desmoplastic Spitz) nevi. They are characterized by epithelioid or spindled
ACCEPTED MANUSCRIPT melanocytes showing prominent surrounding stromal sclerosis. Discussed below are the features of conventional nevi with prominent stromal desmoplasia as they show many overlapping features with desmoplastic melanoma (10). These rare tumors affect adults with a wide age range and a female predilection. A wide
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range of anatomical locations is involved including the trunk, extremities and the head and neck area (10, 11).
Histologically, they are well-demarcated but may show lesional asymmetry (Fig 3a). The dermis is involved but extension into superficial subcutaneous adipose tissue may also be seen. The tumors are composed of epithelioid to spindled melanocytes, arranged singly or
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in small nests or short fascicles in a densely sclerotic stroma (Fig 3b). Peripheral lymphoid
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aggregates may also be noted (12). Cytological atypia is mild, and mitoses are usually not identified (Fig 3c). A pre-existing common acquired, or congenital nevus may be seen in the
the tumor (Fig 3e).
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superficial aspect (Fig 3d) and more nevic melanocytic aggregates may be found throughout
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By immunohistochemistry, the melanocytes express S100, Melan A and P16, but they are
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negative for HMB45 (12) (Fig 3f and g). No chromosomal abnormalities have been detected by conventional four-probe melanoma
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FISH analysis (13).
In view of the spindle cell differentiation in a desmoplastic stroma showing peripheral lymphoid aggregates, the distinction from desmoplastic melanoma is challenging. Helpful histological features include the preserved lesional circumscription with lack of an infiltrative growth pattern and the identification of a background of a conventional nevus. Further supporting findings are the retained melan A and P16 expression on immunohistochemistry and negative results by conventional melanoma FISH.
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1.4. Plaque-type blue nevus Blue nevi are a distinctive variant of melanocytic nevi characterized by the combination of uniform ovoid melanocytes and distinctive pigmented dendritic melanocytes. Their
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morphologic spectrum is wide. They may be pauci- or hypocellular and show stromal sclerosis. The large plaque-type variant is rare but shows significant overlap with desmoplastic melanoma.
Plaque-type blue nevi present as large hyperpigmented dark brown to bluish plaques, measuring multiple centimeters in diameter. They affect all ages and anatomic sites and
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there is no marked gender predilection. The nevi are entirely benign although rare
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malignant transformation (malignant blue nevus) has been reported (14, 15). On histology, the tumors are characterized by a diffuse growth in dermis with extension into
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deeper tissues including subcutaneous adipose tissue, skeletal muscle and fascia or galea (Fig 4a and b). They are composed of a proliferation of ovoid spindle cells arranged singly or
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in nests and short fascicles. The cellularity is variable, ranging from hypercellular areas
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resembling cellular blue nevi to hypocellular areas with variable dermal sclerosis (Fig 4c and d). Admixed are pigmented dendritic melanocytes (Fig 4e). Cytological atypia is mild at most
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and mitotic figures are rare. By immunohistochemistry, melanocytes express SOX10, S100, melan A and HMB45 as observed in other types of blue nevi (Fig. 4f, g and h). Genetically, blue nevi are characterized by mutations in GNAQ or GNA11 (16). The tumors are readily mistaken for desmoplastic melanoma in view of their large size and plaque-like growth with a diffuse and infiltrative architecture and invasion of deeper structures. Reassuring features are the degree of clinically apparent pigmentation, the bland and uniform cytomorphology and the presence of the characteristic pigmented dendritic
ACCEPTED MANUSCRIPT melanocytes. In contrast to desmoplastic melanoma, the expression of both melan A and HMB45 is retained in the spindled melanocytes. Finally, the demonstration of mutations in GNAQ or GNA11 is further support of a diagnosis of plaque type blue nevus.
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2. Acral melanoctic tumors 2.1. Acral lentiginous melanoma
In contrast to melanoma on sun-exposed sites, acral melanoma affects all ethnic groups with a similar incidence. It shows a predilection for the palms and soles, particularly the heel, of elderly adults with a median of 70 years and a slight female predominance (17, 18).
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The tumors are irregular and variably pigmented and may become nodular, deeply
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pigmented and ulcerated with increasing tumor thickness. The overall prognosis is poor as tumors frequently present with advanced tumor thickness (19).
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Histologically, the tumors are poorly demarcated and asymmetrical with an extensive in situ component characterized by a predominantly lentiginous growth of atypical,
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hyperchromatic melanocytes and Pagetoid spread (Fig. 5a, b and c). The involved epidermis
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is typically acanthotic rather than atrophic, and the findings may be subtle and diagnostically challenging, especially in early tumors and with increasing distance from the
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invasive component. The invasive component is characterized by a sheet-like growth of epithelioid or spindled melanocytes showing significant cytologic atypia (Fig.5.d). There is lack of maturation and increased mitotic activity (Fig. 5e). Deep tumors may show tumor necrosis and lymphovascular invasion. The epidermis overlying the invasive component may be atrophic or ulcerated (Fig. 5f). By immunohistochemistry, S100, Melan A and HMB45 expression is preserved in the junctional component (Fig. 5g). In addition to S100 and melan A, HMB45 expression may
ACCEPTED MANUSCRIPT also be seen in the dermal aspect, especially in deeply invasive tumors. P16 and P21 expression a focal finding, observed in less than 25% of dermal melanocytes. Genetically, mutations in BRAF, NRAS, NF1, GNAQ and KIT are most frequently seen (20). Especially, in the early stages acral lentiginous melanoma is difficult to separate reliably
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from acral nevi and acral Spitz nevi which are discussed in detail below (21, 22).
2.2. Atypical acral nevus
Atypical acral nevi are not infrequently encountered. They belong to the group of special site nevi and are characterized by atypical architectural features.
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Clinically, they present as evenly pigmented macules, measuring around 0.5 cm in diameter,
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often with irregular borders, and a predilection for the feet. Although the age range at presentation is wide, they most often present in adults in their forties and there is a female
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predominance. Their behavior is entirely benign (23, 24). Histologically, atypical acral nevi are circumscribed and symmetrical (Fig. 6a). The junctional
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component is lentiginous and nested and composed of small epithelioid melanocytes,
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containing variable amounts of coarse melanin pigment (Fig. 6b). Cytological atypia is not seen. The junctional melanocytic nests are of varying shapes and sizes and they are typically
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located at the tip of the slightly elongated rete ridges. In addition, there may be suprabasilar extension of single melanocytes with extension into the granular cell layer (Fig.6c). When present, these nevi have also been referred to as ‘melanocytic acral nevi with intraepidermal ascent of cells’ (25). Importantly, the intraepidermal ascent of melanocytes is a focal finding confined to the center of the lesion, and it is not encountered at the lesional edges. An additional finding is the presence of columns of melanin pigment in the
ACCEPTED MANUSCRIPT stratum corneum (26, 27). The dermal component is characterized by lack of cytologic atypia, preserved maturation with depth and lack of mitotic activity (Fig. 6d). By immunohistochemistry, melanocytes express S100 and melan A while HMB45 staining is confined to the junctional and superficial dermal components (Fig. 6e). P16 expression in
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the dermal component is strong and diffuse while P21 staining is absent (22). Similar to acral lentiginous melanoma, the most common mutations in atypical acral nevi are seen in BRAF, NRAS, NF1, GNAQ and KIT (20).
Because of the architectural disorder with suprabasilar ascent of melanocytes, atypica acral nevi are easily mistaken for acral lentiginous melanoma. Helpful discriminatory morphologic
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features are the small size, the preserved lesional circumscription, the lack of cytological
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atypia, confinement of Pagetoid spread to the center of the lesion and the preserved maturation with depth of the dermal component. The distinctive immunohistochemical
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2.3. Acral Spitz nevus
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P16+/P21- expression profile further supports the diagnosis of an atypical acral nevi.
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Spitz nevi on acral sites create a particular diagnostic challenge both clinically and histologically, and they are difficult to reliably separate from acral melanoma due to the
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atypical architectural features of nevi on acral sites and the Spitzoid cytologic features. Although the age range is wide (4-61 years) they show a marked predilection for young adults in their twenties and females are more commonly affected than males (22). The feet are twice as frequently involved as the hands. Clinically, the nevi present as asymmetrical pigmented lesions with irregular borders measuring around 0.5 cm in diameter. Larger tumors of up to 1.5 cm have also been reported and melanoma is often suspected clinically. The clinical behavior of acral Spitz nevi is benign (22).
ACCEPTED MANUSCRIPT Histologically, the tumors may show a degree of asymmetry, but they are typically wellcircumscribed (Fig. 7a). The junctional component is broad and, if present, extends significantly beyond the dermal aspect. It shows a lentiginous and nested growth pattern and florid Pagetoid spread is seen in the center of the lesion (Fig. 7b, c and d). The
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melanocytes are epithelioid or spindled with abundant cytoplasm and vesicular nuclei containing small eosinophilic nucleoli. Additional findings are cleft artefacts surrounding the junctional melanocytic nests, transepidermal elimination of nests and ‘Kamino body’ formation. The surrounding epidermis is hyperplastic and epidermal atrophy is rare. The dermal component is cytologically similar to the junctional. Maturation is preserved with
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depth and dermal mitoses are rarely encountered and should be limited to the superficial
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dermal aspect (Fig. 7e). Furthermore, nuclear pleomorphism is not a feature. A focal chronic inflammatory cell infiltrate with evidence of active dermal regression may be seen in a
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subset of tumors.
By immunohistochemistry, melanocytes express S100 and melan A but expression of
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HMB45 is confined to the junctional and superficial dermal component (Fig. 7f). In addition,
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strong and diffuse expression of both P16 and P21 is found in the dermal component (22). The degree of architectural disorder in combination with the Spitzoid cytomorphology
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makes the separation from acral lentiginous melanoma challenging in daily practice. Helpful features include the young patient age, the lesional circumscription, the recognition of the Spitzoid cytomorphology and the preserved maturation with depth of the dermal component with lack of dermal mitotic activity. By immunohistochemistry the P16+/P21+ expression profile provides further support for the diagnosis of an acral Spitz nevus.
3. Vulvar melanocytic tumors
ACCEPTED MANUSCRIPT 3.1. Vulvar melanoma Vulvar melanoma presents as large (around 2 cm), ill-defined, variably pigmented and often ulcerated tumors with a predilection for the mucosal aspect of the vulva. The labia majora are most frequently involved. The majority of tumors affects postmenopausal women in the
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60s and 70s without ethnic predilections (28-30). The overall prognosis is poor as the majority of tumors are deeply invasive at the time of presentation (30-33).
The most common morphologic subtype of melanoma presenting on the vulva is mucosal lentiginous followed by nodular melanoma. The mucosal lentiginous melanomas are broad, ill-defined and asymmetric (Fig 8a). The junctional component is extensive with a diffuse
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and contiguous growth of single melanocytes (Fig 8b and c). The overall features of the in
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situ component may be subtle and cytological atypia is often minimal, especially with increasing distance from the invasive tumor. The invasive component is frequently deep at
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presentation and characterized by a sheet-like growth of severely atypical epithelioid or spindled tumor cells showing brisk mitotic activity and lack of maturation with depth (Fig
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8d). Ulceration, tumor necrosis and lymphovascular invasion may also be present (Fig 8e).
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Tumor cells express S100 and melan A, which also highlight the extent of the in situ component (Fig 8c). HMB45 expression may also be seen in the invasive tumor.
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Genetically, mucosal melanomas show mutations in the SF3B1 and KIT genes but not BRAF or NRAS (34, 35).
The differential diagnosis of vulvar melanoma includes both melanocytic and nonmelanocytic tumors. Pigmented seborrheic keratosis, squamous cell carcinoma and genital melanosis can be easily mistaken for melanoma clinically. They are however squamous proliferations and lack expression of S100 and melan A. Extramammary Paget disease very
ACCEPTED MANUSCRIPT closely resembles melanoma histologically but the tumor cells express cytokeratins and they are S100 and melan A negative. Atypical gential nevus is discussed in more detail below.
3.2. Atypical genital nevus
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Atypical gential nevi belong to the group of special site nevi. They show significant architectural as well as cytological atypia and are readily mistaken for invasive melanoma (36).
They are concerning clinically as they present as large (up to 2 cm), heavily pigmented lesions with irregular borders. There is a strong predilection for young females with a
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median age of 26 years. Both mucosal and cutaneous surfaces of the vulva are affected,
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and they may also be seen in the perineum and the mons pubis. Their behavior is entirely benign with rare local recurrence only if incompletely excised (36, 37).
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Histologically, the tumors are large and often show a prominent, dermal component (Fig 9a). The overlying junctional component is composed of variably shaped and irregularly
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distributed melanocytic nests of varying size (Fig 9b). A lentiginous growth pattern is a
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minor feature but Pagetoid spread into the granular cell layer and growth along skin adnexal structures may be seen (Fig 9c). There is lack of a shoulder and the junctional component
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does not extends significantly beyond the dermal aspect. The junctional melanocytes are epithelioid and contain abundant cytoplasm with coarse melanin. Melanin pigmentation may be prominent and seen in melanocytes and histiocytes with an uneven distribution across the lesion (Fig 9d). Moderate to severe cytological atypia is present in the majority of cases (Fig 9e). It is confluent rather than scattered may also involve the superficial dermal aspect. Maturation with depth is preserved and there is no cytological atypia in the deeper dermal reaches (Fig 9f). Similarly, mitotic activity is frequently present but confined to the
ACCEPTED MANUSCRIPT superficial dermal melanocytic component. An additional finding is the presence of bandlike areas of fibrosis in the superficial dermis (Fig 9g). Immunohistochemically, S100 and melan A are expressed strongly and diffusely while HMB45 expression is confined to the junctional and superficial dermal aspects (Fig 9h and i).
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BRAF mutations are commonly seen in atypical genital nevi (34, 35). Helpful findings in favor of atypical genital nevus over lentiginous mucosal melanoma are the younger age group at presentation, the lesional circumscription and symmetry, lack of Pagetoid spread at the periphery of the tumor, preserved maturation with depth of the dermal compartment, lack of mitotic activity and cytological atypia in the deeper dermal
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compartments. Identification of BRAF mutations is a further helpful feature.
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Conclusion
Although reliable differentiation of the above-mentioned variants of melanocytic nevi from
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melanoma is challenging, attention to the subtle morphological differentiating features, interpretation of the findings in the appropriate clinical context and knowledge of the
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immunohistochemical profile and molecular genetics allows an accurate and confident
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diagnosis and prevents against overtreatment.
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histological study according to the plane of section. Am J Surg Pathol. 1999;23:283-287. 28. Moxley KM, Fader AN, Rose PG, et al. Malignant melanoma of the vulva: an extension of cutaneous melanoma? Gynecol Oncol. 2011;122:612-617. 29. Stang A, Streller B, Eisinger B, et al. Population-based incidence rates of malignant melanoma of the vulva in Germany. Gynecol Oncol. 2005;96:216-221.
ACCEPTED MANUSCRIPT 30. Weinstock MA. Malignant melanoma of the vulva and vagina in the United States: patterns of incidence and population-based estimates of survival. Am J Obstet Gynecol. 1994;171:1225-1230. 31. Raspagliesi F, Ditto A, Paladini D, et al. Prognostic indicators in melanoma of the vulva.
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Ann Surg Oncol. 2000;7:738-742. 32. Ragnarsson-Olding BK, Nilsson BR, Kanter-Lewensohn LR, et al. Malignant melanoma of the vulva in a nationwide, 25-year study of 219 Swedish females: predictors of survival. Cancer. 1999;86:1285-1293.
patients. Cancer. 1996;78:2353-2358.
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33. Raber G, Mempel V, Jackisch C, et al. Malignant melanoma of the vulva. Report of 89
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34. Tseng D, Kim J, Warrick A, et al. Oncogenic mutations in melanomas and benign melanocytic nevi of the female genital tract. J Am Acad Dermatol. 2014;71:229-236.
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35. Yelamos O, Merkel EA, Sholl LM, et al. Nonoverlapping Clinical and Mutational Patterns in Melanomas from the Female Genital Tract and Atypical Genital Nevi. J Invest
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Dermatol. 2016;136:1858-1865.
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36. Clark WH, Jr., Hood AF, Tucker MA, et al. Atypical melanocytic nevi of the genital type with a discussion of reciprocal parenchymal-stromal interactions in the biology of
37.
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neoplasia. Hum Pathol. 1998;29:S1-24. Gleason BC, Hirsch MS, Nucci MR, et al. Atypical genital nevi. A clinicopathologic
analysis of 56 cases. Am J Surg Pathol. 2008;32:51-57.
ACCEPTED MANUSCRIPT Figure legends Fig. 1. Desmoplastic melanoma. This poorly circumscribed, dermal based tumor shows a diffusely infiltrative growth into underlying subcutaneous adipose tissue. It is characterized by prominent sclerosis and multiple peripheral lymphoid aggregates (a). No junctional
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component is present (b) and the hallmark of the tumor is a paucicellular spindle cell proliferation in a desmoplastic stroma (c). The tumor cells are hyperchromatic but there is little cytologic atypia or nuclear pleomorphism (d). Immunohistochemistry for S100 highlights the diffusely infiltrative tumor growth (e).
Fig. 2. Nevi with perineurial differentiation. This tumor is dermal based tumor is relatively
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circumscribed but shows extension into subcutis (a). It has a biphasic growth pattern with a
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superficial nevic component and a deeper spindle cell proliferation (b). The spindle cells are bland and uniform and they are arranged in short interesting fascicles in a variably myxoid
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or desmoplastic stroma (c). In areas, the spindled tumor cells contain wavy and somewhat hyperchromatic nuclei on a densely collagenous matrix with a somewhat storiform growth
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(d). Small hypocellular nodules (e) and peripheral lymphoid aggregates (f) are further
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characteristic findings. The spindle cells compartment is composed of S100-positive (red)
(h).
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and a second population of EMA-positive (brown) (g) cells showing co-expression of glut1
Fig. 3: Desmoplastic nevus. The tumor appears nodular, well-demarcated but asymmetrical (a), composed of epithelioid and spindled melanocytes in a desmoplastic stroma (b). Cytological atypia is not a feature (c). A pre-existing banal dermal nevus is seen in the superficial aspect of the tumor (d), and small nevic nests may be seen throughout the lesion (e). By immunohistochemistry tumors cells express melan a (f) and P16 strongly and diffusely (g).
ACCEPTED MANUSCRIPT Fig. 4. Plaque-type blue nevus. The tumors are large and characterized by a diffuse growth with involvement of subcutis (a) and underlying fascia (b). They are composed of spindle cells arranged singly in a desmoplastic stroma (c) or in intersecting fascicles (d). At least focally, there is a background of a recognizable blue nevus showing pigmented dendritic
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cells (e). Expression of S100 is variable (f) but there is strong and diffuse staining for melan a (g) and HMB45 (h).
Fig. 5. Acral lentiginous melanoma. These large tumors are ill-defined and asymmetrical with ulceration overlying the invasive tumor and an extensive adjacent in situ component (a). The in situ melanoma shows a diffuse and predominantly lentiginous growth pattern (b).
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It is composed of epithelioid melanocytes with hyperchromatic nuclei and a contiguous
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growth with focal Pagetoid spread (c). The invasive tumor is composed of pleomorphic epithelioid melanocytes with brisk mitotic activity and a sheet-like growth pattern (d and e).
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The overlying epidermis is atrophic (f). Immunohistochemistry for melan a highlights the extensive in situ melanoma (g).
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Fig. 6. Acral nevus. The nevi are small and circumscribed (a) with a lentiginous and nested
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growth pattern (b). There is focal Pagetoid spread but cytologic atypia is not seen (c). The dermal component is bland with preserved maturation with depth (d).
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Immunohistochemistry for melan a highlights the lesional circumscription and symmetry (e). Fig. 7. Acral Spitz nevus. The nevi are large but well-circumscribed and relatively symmetrical (a). Their junctional component is lentiginous (b) and nested and composed of epithelioid melanocytes with a Spitzoid cytomorphology (c). Also note the epidermal hyperplasia and the cleft artefact surrounding the junctional melanocytic nests (c). Pagetoid spread is noted in the center of the lesion (d). Rare mitoses may be seen in the superficial dermal component (e). Immunohistochemistry for melan a highlights Pagetoid spread (f).
ACCEPTED MANUSCRIPT Fig. 8. Vulvar mucosal lentiginous melanoma. The tumor is large, poorly-circumscribed and asymmetrical with a deeply invasive component and extensive adjacent in situ melanoma. The in situ component may be subtle (b) and the contiguous growth of melanocytes may be best appreciated on immunohistochemistry for melan a (c). The invasive tumor is composed
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of a sheet-like growth of pleomorphic epithelioid melanocytes (d) with overlying ulceration (e).
Fig. 9. Atypical genital nevus. The tumor is circumscribed but not entirely symmetrical. Also note the prominent dermal component (a). The junctional component is composed of variably sized and shaped melanocytic nests irregularly distributed along the epidermis (b).
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There is extension along hair follicles (c). There is irregular pigment distribution across the
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lesion (d). The junctional melanocytes are epithelioid with a degree of cytological atypia (e). The underlying dermal component is bland with preserved maturation with depth (f). Broad
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bands of dermal fibrosis may be present (g). Immunohistochemistry for melan a highlights
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the lesional circumscription (h) and Pagetoid spread (i).
ACCEPTED MANUSCRIPT
Circumscribed but asymmetric Involvement of superficial subcutis Pre-existing nevus Lack of cytological atypia Poorly- circumscribed and asymmetric Infiltrative growth with deep extension Lack of cytological atypia Pigmented dendritic cells and areas of more conventional blue nevus
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Plaque type blue nevus
Wide anatomic distribution Wide age range F>M Wide anatomic distribution Wide age range M=F
IHC S100+ Melan aHMB45 -
Mutations NF-1 Lack of BRAF
S100+, Melan a-, HMB45and S100,EMA+, GLUT1+ S100+ Melan a+ HMB45-
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Desmoplastic melanocytic nevus
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Melanocytic nevus with perineurial differentiation
Histology Poorly-circumscribed Infiltrative growth with deep extension At least some degree of cytological atypia Circumscribed but asymmetric Involvement of superficial subcutis Pre-existing nevus Lack of cytological atypia
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Desmoplastic melanoma
Clinical Chronically sundamaged skin Elderly M>>F Wide anatomic distribution Adulthood M>F
S100+ (weak) Melan a+ HMB45+
GNAQ GNA11
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Table 1. Salient clinical, histological, immunohistochemical and genetic features if nevi and melanomas with a desmoplastic stroma
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Foot>>hand Young adulthood F>M
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IHC S100+ Melan a+/HMB45 +/P16 focal P21 focal
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Acral Spitz nevus
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Foot>>hand Adulthood F>M
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Atypical acral nevus
Histology Poorly-circumscribed and asymmetric Diffuse growth with extensive junctional component Diffuse Pagetoid spread Cytological atypia of invasive component Lack of maturation with depth Dermal mitoses Ulceration Circumscribed and symmetric Lentiginous and nested Pagetoid spread in center of lesion Lack of cytological atypia Maturation with depth Lack of dermal mitoses Circumscribed but rarely asymmetric Lentiginous and nested Pagetoid spread in center of lesion Spitzoid epithelioid or spindled cytology Rare superficial dermal mitoses
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Acral lentiginous melanoma
Clinical Sole of foot, heel>>hand Elderly F>M
S100+ Melan a+ HMB45P16+ P21-
Mutations BRAF NRAS NF1 GNAQ KIT
BRAF NRAS NF1 GNAQ KIT
S100+ Melan a+ HMB45P16+ P21+
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Table 2. Salient clinical, histological, immunohistochemical and genetic features if nevi and melanomas at acral sites
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Mucosal lentiginous melanoma
Labia majora and perineum Young adulthood
IHC S100+ Melan a+/HMB45 +/-
S100+ Melan a+ HMB45-
Mutations SF3B1 KIT
BRAF
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Atypical gential nevus
Histology Poorly-circumscribed and asymmetric Diffuse growth with extensive junctional component Diffuse Pagetoid spread Cytological atypia of invasive component Lack of maturation with depth Dermal mitoses Ulceration Circumscribed but asymmetric Irregular distribution of junctional nests Pagetoid spread in center of lesion Junctional and superficial dermal cytological atypia Maturation with depth Superficial dermal mitoses
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Clinical Labia majora Mucosa=cutaneous aspect Elderly
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Table 3. Salient clinical, histological, immunohistochemical and genetic features if nevi and melanomas at vulvar sites
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Highlights x Melanocytic tumors pose a significant diagnostic challenge x Broad morphologic spectrum of melanocytic tumors x Overlapping morphological features of benign melanocytic nevi and melanoma x Definitive diagnosis requires awareness of the morphologic spectrum and the subtle morphologic differentiating features, knowledge of immunohistochemical profile and molecular aberrations, and interpretation in the appropriate clinical context
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ACCEPTED MANUSCRIPT Melanocytic Lesions – Staying Out of Trouble
Thomas Brenn, MD, PhD, FRCPatha, b
of Pathology & Laboratory Medicine and Medicine and
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aDepartments
The Arnie Charbonneau Cancer Institute Cumming School of Medicine University of Calgary Diagnostic and Scientific Centre
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9 – 3535 Research Rd NW
Canada
of Pathology
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bDepartment
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Email: [email protected]
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University of Edinburgh
Western General Hospital
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Crewe Rd
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Calgary, AB, T2L 2K8
Edinburgh, EH4 2XU United Kingdom
Declarations of interest: none
ACCEPTED MANUSCRIPT Abstract The diagnosis of melanocytic tumors is notoriously difficult and represents one of the most challenging areas in surgical pathology associated with significant risk for litigation. One reason is the wide morphologic spectrum of melanocytic tumors and the fact that many
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histological features are shared by both benign melanocytic nevi and melanoma. Awareness of the many morphologic variations and variants of nevi and melanoma, their clinical setting, immunohistochemical phenotype and genetic profile is necessary for the correct diagnosis. The article discusses the features of three variants of melanoma, desmoplastic, acral lentiginous and vulvar mucosal lentiginous melanoma, and their benign melanocytic
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mimics with emphasis on distinguishing features.
Keywords
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Melanoma, Nevi, Melanocytic, Immunohistochemistry, Genetics, Diagnosis
ACCEPTED MANUSCRIPT Introduction The correct diagnosis of melanocytic lesions is one of the biggest challenges in dermatopathology and one of the most frequent causes for litigation against Pathologists. There are multiple explanations for this. Some relate to the accuracy or lack of important
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clinical data such as age, gender, anatomical location, clinical presentation and level of clinical suspicion. Others concern the type of biopsy. There is a trend to smaller and more superficial biopsies, which may not be representative of the entire lesion. Finally, the morphologic spectrum of melanocytic tumors is wide with overlapping features of melanoma and benign nevi creating significant diagnostic pitfalls. The following text
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contains a discussion of some of the variants of benign melanocytic nevi which may be
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difficult to separate from invasive melanoma. Although some of these nevi are rare their
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recognition is important to avoid overdiagnosis as melanoma.
1. Melanocytic tumors with stromal desmoplasia
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1.1 Desmoplastic melanoma
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Desmoplastic melanoma is an unusual but distinctive variant of invasive melanoma linked to chronic UV exposure. It shows specific clinical, histological, immunohistochemical and
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genetic features and behavior. Desmoplastic melanoma is a frequent cause for misdiagnosis, especially on partial samples, with a broad differential diagnosis. Clinically, desmoplastic melanoma shows a strong predilection for chronically sun-damaged skin of the head and neck, particularly the scalp, of elderly males. It presents as slowly growing flesh colored nodules and plaques. Clinically visible pigment is seen in a minority of tumors (1).
ACCEPTED MANUSCRIPT Due to its infiltrative growth pattern and neurotropism desmoplastic melanoma shows high local recurrence rates. The overall outcome of ‘pure desmoplastic melanoma’ appears to be favorable compared to conventional melanoma when adjusted for tumor thickness. The tumors also show reduced rates of metastasis to locoregional lymph nodes (2). In contrast,
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the outcome and lymph node metastatic rates of ‘mixed desmoplastic melanoma’ are similar to conventional melanoma.
Histologically, the tumors are poorly circumscribed and asymmetrical with a plaque-like growth pattern in the dermis and a diffusely infiltrative growth into underlying subcutaneous adipose tissue, skeletal muscle or fascia. On low-power inspection the tumors
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are broad, ill-defined and asymmetric (Fig 1a). So-called ‘pure desmoplastic melanoma’ is
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composed of spindle cells arranged singly, in small nests or short fascicles in a desmoplastic stroma (Fig 1b and c). The tumors are paucicellular although the overall cellularity is
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variable. Cytological atypia is usually mild with nuclear hyperchromasia, and the mitotic activity is low (Fig 1d). Peripheral lymphoid aggregates are present and are a good low-
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power diagnostic clue (Fig 1a). Perineurial infiltration is frequently identified. There may be
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an atypical lentiginous melanocytic proliferation involving the overlying epidermis, in some instances amounting to lentigo maligna. Classic melanoma in situ is only rarely encountered.
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A subset of tumors, so-called ‘mixed desmoplastic melanoma’, is characterized by an additional second, more conventional epithelioid or spindled invasive dermal component, characterized by a cellular and sheet-like growth and pronounced cytologic atypia (3, 4). By immunohistochemistry, ‘pure desmoplastic melanoma’ expresses S100, SOX10 and P75 consistently but they are typically negative for melan A, Mart1 and HMB45 (Fig 1e). P16 is expressed in a subset of desmoplastic melanomas (5).
ACCEPTED MANUSCRIPT Genetically, desmoplastic melanomas show frequent mutations in the NF-1 gene but mutations in BRAF, NRAS, GNAQ, GNA11 or KIT are absent (6, 7). The differential diagnosis of desmoplastic melanoma is wide and includes melanocytic and non-melanocytic tumors. Non-melanocytic entities, such as spindle cell squamous cell
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carcinoma, atypical fibroxanthoma, pleomorphic dermal sarcoma, fibrous histiocytoma, dermatofibrosarcoma protubertans and scars can readily be separated by the absence of S100 or SOX10 expression. In contrast the distinction from neurofibroma, especially plaquetype neurofibroma, is more challenging as both tumors share S100 and SOX10 expression. Lack of cytological atypia, absence of the desmoplastic stroma and strong and diffuse CD34
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expression are helpful morphologic and immunohistochemical clues. Furthermore, diffuse
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neurofibroma occurs in young adulthood and does not favor chronically sun-damaged skin. A number of benign melanocytic nevi that may be readily mistaken for desmoplastic
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melanoma are discussed in more detail.
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1.2. Melanocytic nevi with perineurial differentiation (perineuriomatous nevi)
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Melanocytic nevi with perineurial differentiation are a rare but histologically distinctive and potentially under-recognized variant of acquired nevi showing extensive neurotization with
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perineurial differentiation. They are easily mistaken for desmoplastic melanoma on histology (8, 9).
Clinically, they present in adulthood with a male predominance and wide anatomical distribution affecting the trunk, the extremities, the neck and scalp. Often there is a history of a longstanding pigmented lesion with recent change. Their behavior is entirely benign (8, 9).
ACCEPTED MANUSCRIPT Histologically, the tumors are relatively circumscribed but asymmetric (Fig. 2a). They involve the entire dermis and may extend into the superficial subcutaneous adipose tissue. Their growth pattern is biphasic with a common acquired compound or dermal nevus component in the superficial aspect (Fig. 2b). In the deeper reaches, the tumor is composed of uniform
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and bland spindle cells arranged in intersecting fascicles in a variably myxoid and desmoplastic stroma (Fig. 2c). In areas, the spindle cells show slender, wavy nuclei and long, bipolar cytoplasmic processes reminiscent of perineurial cells (Fig. 2d). There is no cytologic atypia or increased mitotic activity. Additional characteristic features include intratumoral hypocellular sclerotic nodules and peripheral lymphoid aggregates (2e and f).
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By immunohistochemistry, the conventional nevic component expresses S100 and melan A
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strongly and diffusely, while HMB45 expression is confined to the junctional and superficial dermal aspect only. The deeper spindle cell component is comprised of and admixture of
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S100-positive, melan A-negative cells and S100-, melan A-negative cells showing expression of the perineurial markers EMA, glut1 and claudin1(8) (Fig. 2g and h).
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Separation from desmoplastic melanoma is challenging on histology. Helpful features
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include the overall circumscription, the lack of cytological atypia, the presence of the distinctive hypocellular nodules and the presence of a common acquired compound or
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dermal nevus in the superficial aspects of the tumor. The clinical presentation further aids the distinction. The tumors have a longstanding history and they show no predilection for chronically sun-damaged skin of the elderly.
1.3. Desmoplastic melanocytic nevi A desmoplastic stroma may be seen in different subtypes of nevi including blue (sclerosing blue) and Spitz (desmoplastic Spitz) nevi. They are characterized by epithelioid or spindled
ACCEPTED MANUSCRIPT melanocytes showing prominent surrounding stromal sclerosis. Discussed below are the features of conventional nevi with prominent stromal desmoplasia as they show many overlapping features with desmoplastic melanoma (10). These rare tumors affect adults with a wide age range and a female predilection. A wide
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range of anatomical locations is involved including the trunk, extremities and the head and neck area (10, 11).
Histologically, they are well-demarcated but may show lesional asymmetry (Fig 3a). The dermis is involved but extension into superficial subcutaneous adipose tissue may also be seen. The tumors are composed of epithelioid to spindled melanocytes, arranged singly or
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in small nests or short fascicles in a densely sclerotic stroma (Fig 3b). Peripheral lymphoid
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aggregates may also be noted (12). Cytological atypia is mild, and mitoses are usually not identified (Fig 3c). A pre-existing common acquired, or congenital nevus may be seen in the
the tumor (Fig 3e).
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superficial aspect (Fig 3d) and more nevic melanocytic aggregates may be found throughout
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By immunohistochemistry, the melanocytes express S100, Melan A and P16, but they are
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negative for HMB45 (12) (Fig 3f and g). No chromosomal abnormalities have been detected by conventional four-probe melanoma
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FISH analysis (13).
In view of the spindle cell differentiation in a desmoplastic stroma showing peripheral lymphoid aggregates, the distinction from desmoplastic melanoma is challenging. Helpful histological features include the preserved lesional circumscription with lack of an infiltrative growth pattern and the identification of a background of a conventional nevus. Further supporting findings are the retained melan A and P16 expression on immunohistochemistry and negative results by conventional melanoma FISH.
ACCEPTED MANUSCRIPT
1.4. Plaque-type blue nevus Blue nevi are a distinctive variant of melanocytic nevi characterized by the combination of uniform ovoid melanocytes and distinctive pigmented dendritic melanocytes. Their
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morphologic spectrum is wide. They may be pauci- or hypocellular and show stromal sclerosis. The large plaque-type variant is rare but shows significant overlap with desmoplastic melanoma.
Plaque-type blue nevi present as large hyperpigmented dark brown to bluish plaques, measuring multiple centimeters in diameter. They affect all ages and anatomic sites and
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there is no marked gender predilection. The nevi are entirely benign although rare
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malignant transformation (malignant blue nevus) has been reported (14, 15). On histology, the tumors are characterized by a diffuse growth in dermis with extension into
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deeper tissues including subcutaneous adipose tissue, skeletal muscle and fascia or galea (Fig 4a and b). They are composed of a proliferation of ovoid spindle cells arranged singly or
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in nests and short fascicles. The cellularity is variable, ranging from hypercellular areas
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resembling cellular blue nevi to hypocellular areas with variable dermal sclerosis (Fig 4c and d). Admixed are pigmented dendritic melanocytes (Fig 4e). Cytological atypia is mild at most
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and mitotic figures are rare. By immunohistochemistry, melanocytes express SOX10, S100, melan A and HMB45 as observed in other types of blue nevi (Fig. 4f, g and h). Genetically, blue nevi are characterized by mutations in GNAQ or GNA11 (16). The tumors are readily mistaken for desmoplastic melanoma in view of their large size and plaque-like growth with a diffuse and infiltrative architecture and invasion of deeper structures. Reassuring features are the degree of clinically apparent pigmentation, the bland and uniform cytomorphology and the presence of the characteristic pigmented dendritic
ACCEPTED MANUSCRIPT melanocytes. In contrast to desmoplastic melanoma, the expression of both melan A and HMB45 is retained in the spindled melanocytes. Finally, the demonstration of mutations in GNAQ or GNA11 is further support of a diagnosis of plaque type blue nevus.
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2. Acral melanoctic tumors 2.1. Acral lentiginous melanoma
In contrast to melanoma on sun-exposed sites, acral melanoma affects all ethnic groups with a similar incidence. It shows a predilection for the palms and soles, particularly the heel, of elderly adults with a median of 70 years and a slight female predominance (17, 18).
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The tumors are irregular and variably pigmented and may become nodular, deeply
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pigmented and ulcerated with increasing tumor thickness. The overall prognosis is poor as tumors frequently present with advanced tumor thickness (19).
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Histologically, the tumors are poorly demarcated and asymmetrical with an extensive in situ component characterized by a predominantly lentiginous growth of atypical,
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hyperchromatic melanocytes and Pagetoid spread (Fig. 5a, b and c). The involved epidermis
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is typically acanthotic rather than atrophic, and the findings may be subtle and diagnostically challenging, especially in early tumors and with increasing distance from the
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invasive component. The invasive component is characterized by a sheet-like growth of epithelioid or spindled melanocytes showing significant cytologic atypia (Fig.5.d). There is lack of maturation and increased mitotic activity (Fig. 5e). Deep tumors may show tumor necrosis and lymphovascular invasion. The epidermis overlying the invasive component may be atrophic or ulcerated (Fig. 5f). By immunohistochemistry, S100, Melan A and HMB45 expression is preserved in the junctional component (Fig. 5g). In addition to S100 and melan A, HMB45 expression may
ACCEPTED MANUSCRIPT also be seen in the dermal aspect, especially in deeply invasive tumors. P16 and P21 expression a focal finding, observed in less than 25% of dermal melanocytes. Genetically, mutations in BRAF, NRAS, NF1, GNAQ and KIT are most frequently seen (20). Especially, in the early stages acral lentiginous melanoma is difficult to separate reliably
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from acral nevi and acral Spitz nevi which are discussed in detail below (21, 22).
2.2. Atypical acral nevus
Atypical acral nevi are not infrequently encountered. They belong to the group of special site nevi and are characterized by atypical architectural features.
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Clinically, they present as evenly pigmented macules, measuring around 0.5 cm in diameter,
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often with irregular borders, and a predilection for the feet. Although the age range at presentation is wide, they most often present in adults in their forties and there is a female
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predominance. Their behavior is entirely benign (23, 24). Histologically, atypical acral nevi are circumscribed and symmetrical (Fig. 6a). The junctional
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component is lentiginous and nested and composed of small epithelioid melanocytes,
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containing variable amounts of coarse melanin pigment (Fig. 6b). Cytological atypia is not seen. The junctional melanocytic nests are of varying shapes and sizes and they are typically
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located at the tip of the slightly elongated rete ridges. In addition, there may be suprabasilar extension of single melanocytes with extension into the granular cell layer (Fig.6c). When present, these nevi have also been referred to as ‘melanocytic acral nevi with intraepidermal ascent of cells’ (25). Importantly, the intraepidermal ascent of melanocytes is a focal finding confined to the center of the lesion, and it is not encountered at the lesional edges. An additional finding is the presence of columns of melanin pigment in the
ACCEPTED MANUSCRIPT stratum corneum (26, 27). The dermal component is characterized by lack of cytologic atypia, preserved maturation with depth and lack of mitotic activity (Fig. 6d). By immunohistochemistry, melanocytes express S100 and melan A while HMB45 staining is confined to the junctional and superficial dermal components (Fig. 6e). P16 expression in
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the dermal component is strong and diffuse while P21 staining is absent (22). Similar to acral lentiginous melanoma, the most common mutations in atypical acral nevi are seen in BRAF, NRAS, NF1, GNAQ and KIT (20).
Because of the architectural disorder with suprabasilar ascent of melanocytes, atypica acral nevi are easily mistaken for acral lentiginous melanoma. Helpful discriminatory morphologic
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features are the small size, the preserved lesional circumscription, the lack of cytological
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atypia, confinement of Pagetoid spread to the center of the lesion and the preserved maturation with depth of the dermal component. The distinctive immunohistochemical
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2.3. Acral Spitz nevus
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P16+/P21- expression profile further supports the diagnosis of an atypical acral nevi.
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Spitz nevi on acral sites create a particular diagnostic challenge both clinically and histologically, and they are difficult to reliably separate from acral melanoma due to the
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atypical architectural features of nevi on acral sites and the Spitzoid cytologic features. Although the age range is wide (4-61 years) they show a marked predilection for young adults in their twenties and females are more commonly affected than males (22). The feet are twice as frequently involved as the hands. Clinically, the nevi present as asymmetrical pigmented lesions with irregular borders measuring around 0.5 cm in diameter. Larger tumors of up to 1.5 cm have also been reported and melanoma is often suspected clinically. The clinical behavior of acral Spitz nevi is benign (22).
ACCEPTED MANUSCRIPT Histologically, the tumors may show a degree of asymmetry, but they are typically wellcircumscribed (Fig. 7a). The junctional component is broad and, if present, extends significantly beyond the dermal aspect. It shows a lentiginous and nested growth pattern and florid Pagetoid spread is seen in the center of the lesion (Fig. 7b, c and d). The
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melanocytes are epithelioid or spindled with abundant cytoplasm and vesicular nuclei containing small eosinophilic nucleoli. Additional findings are cleft artefacts surrounding the junctional melanocytic nests, transepidermal elimination of nests and ‘Kamino body’ formation. The surrounding epidermis is hyperplastic and epidermal atrophy is rare. The dermal component is cytologically similar to the junctional. Maturation is preserved with
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depth and dermal mitoses are rarely encountered and should be limited to the superficial
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dermal aspect (Fig. 7e). Furthermore, nuclear pleomorphism is not a feature. A focal chronic inflammatory cell infiltrate with evidence of active dermal regression may be seen in a
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subset of tumors.
By immunohistochemistry, melanocytes express S100 and melan A but expression of
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HMB45 is confined to the junctional and superficial dermal component (Fig. 7f). In addition,
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strong and diffuse expression of both P16 and P21 is found in the dermal component (22). The degree of architectural disorder in combination with the Spitzoid cytomorphology
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makes the separation from acral lentiginous melanoma challenging in daily practice. Helpful features include the young patient age, the lesional circumscription, the recognition of the Spitzoid cytomorphology and the preserved maturation with depth of the dermal component with lack of dermal mitotic activity. By immunohistochemistry the P16+/P21+ expression profile provides further support for the diagnosis of an acral Spitz nevus.
3. Vulvar melanocytic tumors
ACCEPTED MANUSCRIPT 3.1. Vulvar melanoma Vulvar melanoma presents as large (around 2 cm), ill-defined, variably pigmented and often ulcerated tumors with a predilection for the mucosal aspect of the vulva. The labia majora are most frequently involved. The majority of tumors affects postmenopausal women in the
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60s and 70s without ethnic predilections (28-30). The overall prognosis is poor as the majority of tumors are deeply invasive at the time of presentation (30-33).
The most common morphologic subtype of melanoma presenting on the vulva is mucosal lentiginous followed by nodular melanoma. The mucosal lentiginous melanomas are broad, ill-defined and asymmetric (Fig 8a). The junctional component is extensive with a diffuse
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and contiguous growth of single melanocytes (Fig 8b and c). The overall features of the in
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situ component may be subtle and cytological atypia is often minimal, especially with increasing distance from the invasive tumor. The invasive component is frequently deep at
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presentation and characterized by a sheet-like growth of severely atypical epithelioid or spindled tumor cells showing brisk mitotic activity and lack of maturation with depth (Fig
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8d). Ulceration, tumor necrosis and lymphovascular invasion may also be present (Fig 8e).
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Tumor cells express S100 and melan A, which also highlight the extent of the in situ component (Fig 8c). HMB45 expression may also be seen in the invasive tumor.
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Genetically, mucosal melanomas show mutations in the SF3B1 and KIT genes but not BRAF or NRAS (34, 35).
The differential diagnosis of vulvar melanoma includes both melanocytic and nonmelanocytic tumors. Pigmented seborrheic keratosis, squamous cell carcinoma and genital melanosis can be easily mistaken for melanoma clinically. They are however squamous proliferations and lack expression of S100 and melan A. Extramammary Paget disease very
ACCEPTED MANUSCRIPT closely resembles melanoma histologically but the tumor cells express cytokeratins and they are S100 and melan A negative. Atypical gential nevus is discussed in more detail below.
3.2. Atypical genital nevus
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Atypical gential nevi belong to the group of special site nevi. They show significant architectural as well as cytological atypia and are readily mistaken for invasive melanoma (36).
They are concerning clinically as they present as large (up to 2 cm), heavily pigmented lesions with irregular borders. There is a strong predilection for young females with a
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median age of 26 years. Both mucosal and cutaneous surfaces of the vulva are affected,
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and they may also be seen in the perineum and the mons pubis. Their behavior is entirely benign with rare local recurrence only if incompletely excised (36, 37).
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Histologically, the tumors are large and often show a prominent, dermal component (Fig 9a). The overlying junctional component is composed of variably shaped and irregularly
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distributed melanocytic nests of varying size (Fig 9b). A lentiginous growth pattern is a
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minor feature but Pagetoid spread into the granular cell layer and growth along skin adnexal structures may be seen (Fig 9c). There is lack of a shoulder and the junctional component
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does not extends significantly beyond the dermal aspect. The junctional melanocytes are epithelioid and contain abundant cytoplasm with coarse melanin. Melanin pigmentation may be prominent and seen in melanocytes and histiocytes with an uneven distribution across the lesion (Fig 9d). Moderate to severe cytological atypia is present in the majority of cases (Fig 9e). It is confluent rather than scattered may also involve the superficial dermal aspect. Maturation with depth is preserved and there is no cytological atypia in the deeper dermal reaches (Fig 9f). Similarly, mitotic activity is frequently present but confined to the
ACCEPTED MANUSCRIPT superficial dermal melanocytic component. An additional finding is the presence of bandlike areas of fibrosis in the superficial dermis (Fig 9g). Immunohistochemically, S100 and melan A are expressed strongly and diffusely while HMB45 expression is confined to the junctional and superficial dermal aspects (Fig 9h and i).
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BRAF mutations are commonly seen in atypical genital nevi (34, 35). Helpful findings in favor of atypical genital nevus over lentiginous mucosal melanoma are the younger age group at presentation, the lesional circumscription and symmetry, lack of Pagetoid spread at the periphery of the tumor, preserved maturation with depth of the dermal compartment, lack of mitotic activity and cytological atypia in the deeper dermal
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compartments. Identification of BRAF mutations is a further helpful feature.
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Conclusion
Although reliable differentiation of the above-mentioned variants of melanocytic nevi from
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melanoma is challenging, attention to the subtle morphological differentiating features, interpretation of the findings in the appropriate clinical context and knowledge of the
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immunohistochemical profile and molecular genetics allows an accurate and confident
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diagnosis and prevents against overtreatment.
ACCEPTED MANUSCRIPT References 1.
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10. Sherrill AM, Crespo G, Prakash AV, et al. Desmoplastic nevus: An entity distinct from spitz nevus and blue nevus. Am J Dermatopathol. 2011;33:35-39.
ACCEPTED MANUSCRIPT 11. Sidiropoulos M, Sholl LM, Obregon R, et al. Desmoplastic nevus of chronically sundamaged skin: an entity to be distinguished from desmoplastic melanoma. Am J Dermatopathol. 2014;36:629-634. 12. Kiuru M, Patel RM, Busam KJ. Desmoplastic melanocytic nevi with lymphocytic
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aggregates. J Cutan Pathol. 2012;39:940-944. 13. Gerami P, Beilfuss B, Haghighat Z, et al. Fluorescence in situ hybridization as an ancillary method for the distinction of desmoplastic melanomas from sclerosing melanocytic nevi. J Cutan Pathol. 2011;38:329-334.
14. Yeh I, Fang Y, Busam KJ. Melanoma arising in a large plaque-type blue nevus with
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15. Busam KJ, Woodruff JM, Erlandson RA, et al. Large plaque-type blue nevus with subcutaneous cellular nodules. Am J Surg Pathol. 2000;24:92-99.
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16. Van Raamsdonk CD, Bezrookove V, Green G, et al. Frequent somatic mutations of GNAQ in uveal melanoma and blue naevi. Nature. 2009;457:599-602.
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18. Kuchelmeister C, Schaumburg-Lever G, Garbe C. Acral cutaneous melanoma in caucasians: clinical features, histopathology and prognosis in 112 patients. Br J Dermatol. 2000;143:275-280. 19. Teramoto Y, Keim U, Gesierich A, et al. Acral lentiginous melanoma: a skin cancer with unfavourable prognostic features. A study of the German central malignant melanoma registry (CMMR) in 2050 patients. Br J Dermatol. 2018;178:443-451.
ACCEPTED MANUSCRIPT 20. Moon KR, Choi YD, Kim JM, et al. Genetic Alterations in Primary Acral Melanoma and Acral Melanocytic Nevus in Korea: Common Mutated Genes Show Distinct Cytomorphological Features. J Invest Dermatol. 2018;138:933-945. 21. Fernandez-Flores A, Cassarino DS. Histopathological diagnosis of acral lentiginous
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melanoma in early stages. Ann Diagn Pathol. 2017;26:64-69. 22. Wiedemeyer K, Guadagno A, Davey J, et al. Acral Spitz Nevi: A Clinicopathologic Study of 50 Cases With Immunohistochemical Analysis of P16 and P21 Expression. Am J Surg Pathol. 2018;42:821-827.
23. Palicka GA, Rhodes AR. Acral melanocytic nevi: prevalence and distribution of gross
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morphologic features in white and black adults. Arch Dermatol. 2010;146:1085-1094.
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24. Boyd AS, Rapini RP. Acral melanocytic neoplasms: a histologic analysis of 158 lesions. J Am Acad Dermatol. 1994;31:740-745.
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25. LeBoit PE. A diagnosis for maniacs. Am J Dermatopathol. 2000;22:556-558. 26. Saida T, Koga H, Goto Y, et al. Characteristic distribution of melanin columns in the
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cornified layer of acquired acral nevus: an important clue for histopathologic
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differentiation from early acral melanoma. Am J Dermatopathol. 2011;33:468-473. 27. Signoretti S, Annessi G, Puddu P, et al. Melanocytic nevi of palms and soles: a
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histological study according to the plane of section. Am J Surg Pathol. 1999;23:283-287. 28. Moxley KM, Fader AN, Rose PG, et al. Malignant melanoma of the vulva: an extension of cutaneous melanoma? Gynecol Oncol. 2011;122:612-617. 29. Stang A, Streller B, Eisinger B, et al. Population-based incidence rates of malignant melanoma of the vulva in Germany. Gynecol Oncol. 2005;96:216-221.
ACCEPTED MANUSCRIPT 30. Weinstock MA. Malignant melanoma of the vulva and vagina in the United States: patterns of incidence and population-based estimates of survival. Am J Obstet Gynecol. 1994;171:1225-1230. 31. Raspagliesi F, Ditto A, Paladini D, et al. Prognostic indicators in melanoma of the vulva.
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Ann Surg Oncol. 2000;7:738-742. 32. Ragnarsson-Olding BK, Nilsson BR, Kanter-Lewensohn LR, et al. Malignant melanoma of the vulva in a nationwide, 25-year study of 219 Swedish females: predictors of survival. Cancer. 1999;86:1285-1293.
patients. Cancer. 1996;78:2353-2358.
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33. Raber G, Mempel V, Jackisch C, et al. Malignant melanoma of the vulva. Report of 89
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34. Tseng D, Kim J, Warrick A, et al. Oncogenic mutations in melanomas and benign melanocytic nevi of the female genital tract. J Am Acad Dermatol. 2014;71:229-236.
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35. Yelamos O, Merkel EA, Sholl LM, et al. Nonoverlapping Clinical and Mutational Patterns in Melanomas from the Female Genital Tract and Atypical Genital Nevi. J Invest
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Dermatol. 2016;136:1858-1865.
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36. Clark WH, Jr., Hood AF, Tucker MA, et al. Atypical melanocytic nevi of the genital type with a discussion of reciprocal parenchymal-stromal interactions in the biology of
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neoplasia. Hum Pathol. 1998;29:S1-24. Gleason BC, Hirsch MS, Nucci MR, et al. Atypical genital nevi. A clinicopathologic
analysis of 56 cases. Am J Surg Pathol. 2008;32:51-57.
ACCEPTED MANUSCRIPT Figure legends Fig. 1. Desmoplastic melanoma. This poorly circumscribed, dermal based tumor shows a diffusely infiltrative growth into underlying subcutaneous adipose tissue. It is characterized by prominent sclerosis and multiple peripheral lymphoid aggregates (a). No junctional
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component is present (b) and the hallmark of the tumor is a paucicellular spindle cell proliferation in a desmoplastic stroma (c). The tumor cells are hyperchromatic but there is little cytologic atypia or nuclear pleomorphism (d). Immunohistochemistry for S100 highlights the diffusely infiltrative tumor growth (e).
Fig. 2. Nevi with perineurial differentiation. This tumor is dermal based tumor is relatively
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circumscribed but shows extension into subcutis (a). It has a biphasic growth pattern with a
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superficial nevic component and a deeper spindle cell proliferation (b). The spindle cells are bland and uniform and they are arranged in short interesting fascicles in a variably myxoid
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or desmoplastic stroma (c). In areas, the spindled tumor cells contain wavy and somewhat hyperchromatic nuclei on a densely collagenous matrix with a somewhat storiform growth
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(d). Small hypocellular nodules (e) and peripheral lymphoid aggregates (f) are further
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characteristic findings. The spindle cells compartment is composed of S100-positive (red)
(h).
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and a second population of EMA-positive (brown) (g) cells showing co-expression of glut1
Fig. 3: Desmoplastic nevus. The tumor appears nodular, well-demarcated but asymmetrical (a), composed of epithelioid and spindled melanocytes in a desmoplastic stroma (b). Cytological atypia is not a feature (c). A pre-existing banal dermal nevus is seen in the superficial aspect of the tumor (d), and small nevic nests may be seen throughout the lesion (e). By immunohistochemistry tumors cells express melan a (f) and P16 strongly and diffusely (g).
ACCEPTED MANUSCRIPT Fig. 4. Plaque-type blue nevus. The tumors are large and characterized by a diffuse growth with involvement of subcutis (a) and underlying fascia (b). They are composed of spindle cells arranged singly in a desmoplastic stroma (c) or in intersecting fascicles (d). At least focally, there is a background of a recognizable blue nevus showing pigmented dendritic
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cells (e). Expression of S100 is variable (f) but there is strong and diffuse staining for melan a (g) and HMB45 (h).
Fig. 5. Acral lentiginous melanoma. These large tumors are ill-defined and asymmetrical with ulceration overlying the invasive tumor and an extensive adjacent in situ component (a). The in situ melanoma shows a diffuse and predominantly lentiginous growth pattern (b).
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It is composed of epithelioid melanocytes with hyperchromatic nuclei and a contiguous
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growth with focal Pagetoid spread (c). The invasive tumor is composed of pleomorphic epithelioid melanocytes with brisk mitotic activity and a sheet-like growth pattern (d and e).
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The overlying epidermis is atrophic (f). Immunohistochemistry for melan a highlights the extensive in situ melanoma (g).
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Fig. 6. Acral nevus. The nevi are small and circumscribed (a) with a lentiginous and nested
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growth pattern (b). There is focal Pagetoid spread but cytologic atypia is not seen (c). The dermal component is bland with preserved maturation with depth (d).
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Immunohistochemistry for melan a highlights the lesional circumscription and symmetry (e). Fig. 7. Acral Spitz nevus. The nevi are large but well-circumscribed and relatively symmetrical (a). Their junctional component is lentiginous (b) and nested and composed of epithelioid melanocytes with a Spitzoid cytomorphology (c). Also note the epidermal hyperplasia and the cleft artefact surrounding the junctional melanocytic nests (c). Pagetoid spread is noted in the center of the lesion (d). Rare mitoses may be seen in the superficial dermal component (e). Immunohistochemistry for melan a highlights Pagetoid spread (f).
ACCEPTED MANUSCRIPT Fig. 8. Vulvar mucosal lentiginous melanoma. The tumor is large, poorly-circumscribed and asymmetrical with a deeply invasive component and extensive adjacent in situ melanoma. The in situ component may be subtle (b) and the contiguous growth of melanocytes may be best appreciated on immunohistochemistry for melan a (c). The invasive tumor is composed
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of a sheet-like growth of pleomorphic epithelioid melanocytes (d) with overlying ulceration (e).
Fig. 9. Atypical genital nevus. The tumor is circumscribed but not entirely symmetrical. Also note the prominent dermal component (a). The junctional component is composed of variably sized and shaped melanocytic nests irregularly distributed along the epidermis (b).
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There is extension along hair follicles (c). There is irregular pigment distribution across the
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lesion (d). The junctional melanocytes are epithelioid with a degree of cytological atypia (e). The underlying dermal component is bland with preserved maturation with depth (f). Broad
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bands of dermal fibrosis may be present (g). Immunohistochemistry for melan a highlights
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the lesional circumscription (h) and Pagetoid spread (i).
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Circumscribed but asymmetric Involvement of superficial subcutis Pre-existing nevus Lack of cytological atypia Poorly- circumscribed and asymmetric Infiltrative growth with deep extension Lack of cytological atypia Pigmented dendritic cells and areas of more conventional blue nevus
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Plaque type blue nevus
Wide anatomic distribution Wide age range F>M Wide anatomic distribution Wide age range M=F
IHC S100+ Melan aHMB45 -
Mutations NF-1 Lack of BRAF
S100+, Melan a-, HMB45and S100,EMA+, GLUT1+ S100+ Melan a+ HMB45-
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Desmoplastic melanocytic nevus
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Melanocytic nevus with perineurial differentiation
Histology Poorly-circumscribed Infiltrative growth with deep extension At least some degree of cytological atypia Circumscribed but asymmetric Involvement of superficial subcutis Pre-existing nevus Lack of cytological atypia
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Desmoplastic melanoma
Clinical Chronically sundamaged skin Elderly M>>F Wide anatomic distribution Adulthood M>F
S100+ (weak) Melan a+ HMB45+
GNAQ GNA11
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Table 1. Salient clinical, histological, immunohistochemical and genetic features if nevi and melanomas with a desmoplastic stroma
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Foot>>hand Young adulthood F>M
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IHC S100+ Melan a+/HMB45 +/P16 focal P21 focal
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Acral Spitz nevus
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Foot>>hand Adulthood F>M
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Atypical acral nevus
Histology Poorly-circumscribed and asymmetric Diffuse growth with extensive junctional component Diffuse Pagetoid spread Cytological atypia of invasive component Lack of maturation with depth Dermal mitoses Ulceration Circumscribed and symmetric Lentiginous and nested Pagetoid spread in center of lesion Lack of cytological atypia Maturation with depth Lack of dermal mitoses Circumscribed but rarely asymmetric Lentiginous and nested Pagetoid spread in center of lesion Spitzoid epithelioid or spindled cytology Rare superficial dermal mitoses
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Acral lentiginous melanoma
Clinical Sole of foot, heel>>hand Elderly F>M
S100+ Melan a+ HMB45P16+ P21-
Mutations BRAF NRAS NF1 GNAQ KIT
BRAF NRAS NF1 GNAQ KIT
S100+ Melan a+ HMB45P16+ P21+
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Table 2. Salient clinical, histological, immunohistochemical and genetic features if nevi and melanomas at acral sites
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Mucosal lentiginous melanoma
Labia majora and perineum Young adulthood
IHC S100+ Melan a+/HMB45 +/-
S100+ Melan a+ HMB45-
Mutations SF3B1 KIT
BRAF
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Atypical gential nevus
Histology Poorly-circumscribed and asymmetric Diffuse growth with extensive junctional component Diffuse Pagetoid spread Cytological atypia of invasive component Lack of maturation with depth Dermal mitoses Ulceration Circumscribed but asymmetric Irregular distribution of junctional nests Pagetoid spread in center of lesion Junctional and superficial dermal cytological atypia Maturation with depth Superficial dermal mitoses
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Clinical Labia majora Mucosa=cutaneous aspect Elderly
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Table 3. Salient clinical, histological, immunohistochemical and genetic features if nevi and melanomas at vulvar sites
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Highlights x Melanocytic tumors pose a significant diagnostic challenge x Broad morphologic spectrum of melanocytic tumors x Overlapping morphological features of benign melanocytic nevi and melanoma x Definitive diagnosis requires awareness of the morphologic spectrum and the subtle morphologic differentiating features, knowledge of immunohistochemical profile and molecular aberrations, and interpretation in the appropriate clinical context
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