Subungual Melanoma with Cartilaginous Differentiation: Molecular Insights

Abstract

Melanoma’s rare capacity to undergo heterologous differentiation can create significant diagnostic challenges. The molecular mechanisms underlying this phenomenon are not well understood. We present an unusual case of subungual melanoma exhibiting extensive cartilaginous differentiation and provide insights into its molecular and cytogenomic features. Histopathologically, the tumor was predominantly composed of nodules of malignant cartilage in association with a smaller population of nested epithelioid to rhabdoid cells. Immunohistochemically, the tumor cells in both components were positive for S100, SOX10, and PRAME, and were negative for Melan-A and HMB-45. Molecular analysis by whole exome DNA sequence did not detect any pathogenic variants in genes commonly implicated in melanoma. Additional analysis by SNP chromosomal microarray revealed a complex genome characterized by numerous chromosomal losses and gains, including homozygous deletion of the CDKN2A locus and a heterozygous deletion of the locus containing EXT2, a tumor suppressor implicated in hereditary multiple osteochondromas and secondary chondrosarcomas. This case underscores the importance of recognizing cartilaginous differentiation as a rare manifestation of melanoma, particularly at subungual sites, and suggests that at least some of these melanomas may be driven by non-canonical molecular pathways.

Introduction

Melanoma is known for its diverse histopathological presentations, in part due to its ability to undergo dramatic phenotypic transformations in the form of heterologous differentiation. While heterologous differentiation is a rare phenomenon in melanoma, it can introduce significant diagnostic challenges when the resultant morphology resembles other primary tumors. Among its various manifestations, melanoma can display fibroblastic/myofibroblastic, Schwannian, perineurial, smooth muscle, rhabdomyosarcomatous, osteocartilaginous, ganglionic, ganglioneuroblastic, neuroendocrine, and epithelial elements¹. The molecular underpinnings associated with heterologous differentiation in melanoma are not well understood. In this report, we highlight an exceptional case of melanoma with extensive cartilaginous differentiation and explore the molecular events associated with this phenotypic transformation.

Case Report

A 79-year-old man presented to dermatology clinic due to a black-blue discoloration beneath his right index finger nail (Figure 1). The discoloration had persisted for six weeks and was not linked to any overt trauma. Physical exam revealed a well demarcated, dark black to purple subungual patch extending from the proximal nail fold, findings suggestive of a subungual hematoma. Over a three-month observation period, the lesion evolved into a subungual nodule with onycholysis and distal subungual debris, raising concerns for a neoplastic process. The differential diagnosis included subungual squamous cell carcinoma, pyogenic granuloma, onychopapilloma, subungual wart, and melanoma. A nail avulsion with matrix biopsy was performed.

Figure 1.

Clinical images of subungual melanoma. (A) The patient presented with dark brown to black discoloration under the right index finger nail. (B, C) The lesion evolved into a subungual nodule over three months. (D) Residual tumor following nail avulsion biopsy. The skin markings outline the planned amputation site.

Histopathologic evaluation of the biopsy revealed cellular nodules of malignant cartilage (Figure 2). Within these nodules, the lacunar spaces contained round to epithelioid cells with enlarged, hyperchromatic, and pleomorphic nuclei. Occasional mitotic figures were noted within the cartilaginous matrix. A smaller component of the tumor manifested as vaguely nested aggregates of epithelioid tumor cells, lacking any cartilaginous matrix. Focally, tumor cells assumed a rhabdoid appearance, characterized by variably eccentric and pleomorphic nuclei and abundant eosinophilic cytoplasm. Prominent nucleoli and occasional nuclear pseudoinclusions were identified. Mitotic figures were conspicuous. The tumor extended to a depth of 3.7 mm.

Figure 2.

Representative histopathological images of tumor. (A) The tumor was predominantly composed of nodules of hyaline cartilage (1x, H&E). (B) The cartilage exhibited hypercellularity and cytological atypia (15x, H&E). (C) A minor component of the tumor was nested with epithelioid cells (35x, H&E). (D) Focally, the tumor cells were pleomorphic with a rhabdoid appearance (32x, H&E).

By immunohistochemistry, the tumor cells within both the nested and cartilaginous components exhibited diffuse positivity for SOX10, S100, and PRAME (Figure 3), while being negative for Melan-A, HMB-45, BRAF-V600E, and NRAS-Q61R. Additionally, the tumor cells exhibited complete loss of p16. Focally, atypical SOX10-positive cells were observed in a separately received fragment of nail epithelium, suspicious for a possible in situ component. Together, the morphologic and immunohistochemical findings supported a diagnosis of subungual melanoma with cartilaginous differentiation.

Figure 3.

Immunohistochemical findings. Both the cartilaginous (A,C,E,G) and nested components (B,D,F,H) of the tumor were strongly and diffusely positive for (A, B) S100, (C,D) SOX10, and (E,F) PRAME. (G, E) p16 expression was absent. All images are 25x magnification.

The patient subsequently underwent a right distal phalanx amputation, revealing extensive residual melanoma. Sentinel lymph node biopsy was performed, which was negative for tumor. Seven months after the initial diagnosis, there was no evidence of recurrence.

Whole exome DNA sequencing was performed and a total of 148 genes, including BRAF, NRAS, NF1, KIT, TERT, GNAQ, and GNA11, were interrogated for pathogenic variants. No predicted pathogenic variants were identified. The tumor had a low mutational burden, with six somatic mutations per megabase (Mb). Additional genomic analysis by single-nucleotide polymorphism-based chromosomal microarray analysis (OncoScan) was performed. This revealed a complex genomic profile characterized by numerous chromosomal losses and gains (Figure 4 and Table 1). Notably, this included a single copy loss of chromosome nine, along with homozygous loss of the CDKN2A locus and a heterozygous loss of EXT2 within chromosome 11p11.2.

Figure 4.

SNP chromosomal microarray plot demonstrating the tumor’s genomic complexity, including numerous copy number losses and gains (summarized in Table 1). The x-axis represents individual chromosomes while the y-axis represents the log2 ratios (top) and B allele frequencies (bottom).

Table 1:

Summary of SNP chromosomal microarray (OncoScan) results

Chromosomal Losses	Chromosomal Gains
1p36.31	5p15.33-p15.31
2 (subclonal loss; median copy number state 1.66 across chr 2)	5p15.2-p13.1 (including RICTOR)
3	6q14.1
5p15.33	6q21
5p15.2	6q27
5p15.1	7q21.13
5q	18p11.32-p11.21
6p11.2-q14.1	19p-q12
6q14.1-q16.3	19q13.11-q13.12
6q21-q27	19q13.31-q13.32
7q34	22q11.1-q13.32
9 (homozygous deletion of 9p21.3, including CDKN2A)
11p15.4
11p12-p11.12 (including EXT2)
14q
15q11.1-q15.2
19q12-q13.11
19q13.2-q13.31
19q13.32-q13.43
21q21.3-q22.3

Discussion

Cartilaginous differentiation in melanoma, while exceedingly rare, has been reported in both primary and metastatic cases.^2–22 This phenomenon can manifest as purely cartilaginous differentiation, as exemplified by our case, or as a composite of cartilaginous and osteoid elements, known as osteocartilaginous differentiation. When present in melanoma, a cartilaginous component generally exhibits increased cellularity, cytological atypia, and/or mitotic activity, supporting transdifferentiation into chrondrosarcoma, rather than a metaplastic change to benign cartilage.^{4–6,8,10–12,15,16,18,20–22} These melanomas typically occur in older patients without a clear sex predilection. They often arise at acral sites, especially subungually, but cases have been documented in diverse anatomic locations, including mucosal sites and skin of chronic and intermittent sun exposure.^3,8,12,16,21 While they are often associated with a high Breslow thickness, the biological behavior of these melanomas, in comparison to conventional melanomas of matched stage, remains to be fully elucidated.

The etiology behind cartilaginous transdifferentiation in melanoma is not completely understood. Subungual cases are often associated with a history of antecedent trauma, suggesting a possible environmental component.⁴ Tissue-specific factors, such as local growth factors, the surrounding extracellular matrix, or peculiarities of the melanocytic stem cell niche, may also contribute. The role of melanoma-inhibiting activity (MIA), a small soluble protein secreted from melanoma cells and chondrocytes, has been proposed as a potential mediator in this morphological transformation, though supporting data are limited.^3,5,7

To our knowledge, only seven reported cases of melanoma with cartilaginous or osteocartilaginous differentiation have been molecularly interrogated (Table 2). Among these, one case harbored an NRAS Q61 mutation and two harbored BRAF V600E mutations.^18,20 The remaining cases, reported recently by Gallagher et al.²¹, were genetically diverse: one harbored loss-of-function mutations in NF1 and CDKN2A, and one had a pathogenic BRAF S467L mutation and co-occurring NRAS V8M mutation. The remaining two cases had mutations of uncertain significance in GNAQ and NF1.

Table 2:

Summary of mutations identified in previously reported cases of melanoma with cartilaginous differentiation

Case	Type of differentiation	Mutations (Tier I)	Mutations (Tier II)	Authors/Reference
1	Cartilaginous	BRAF V600E	N/A	Berro et al.18
2	Cartilaginous	NRAS Q61R	N/A	Sweeney and Royer20
3	Cartilaginous	BRAF S467L	CDKN2A Ala148Thr; CDKN2A Ala20GlyfsTer6; GNAQ NM_002072.3:c.736–5delT [SPLICE]; NF1 G531Ter; NRAS V8M	Gallagher et al.21
4	Cartilaginous	CDKN2A P81L; NF1 R440Ter	BAP1 VLF530KHL; NF1 G4758V; NF1 N2788Y; NFKBIE G34E; PPP6C H200Y; YAE1D1 V4D; YAE1D1 AAAS6GGC	Gallagher et al.21
5	Cartilaginous	None	GNAQ NM_002072.3:c.736–5delT [SPLICE]	Gallagher et al.21
6	Osteocartilaginous	None	NF1 N2788Y; YAE1D1 AAS6GGC	Gallagher et al.21
7	Cartilaginous	BRAF V600E	N/A	Gallagher et al.21

N/A: not available

Our report provides the first comprehensive analysis of both DNA mutations and genome-wide copy number alterations in a melanoma with cartilaginous differentiation. Notably, no predicted pathogenic variants were detected in genes commonly implicated in melanoma, including BRAF, NRAS, NF1, KIT, TERT, GNAQ, and GNA11. This underscores the tumor’s non-canonical genetic makeup and suggests an alternative oncogenic pathway. Moreover, the tumor’s low-mutational burden, at six per Mb, defies the expected profile of a highly mutated melanoma. This aspect of the tumor’s molecular profile could speak to its subungual origins, considering this is a largely sun protected site. Despite the absence of detectable pathogenic variants, chromosomal microarray analysis revealed a structurally complex genome marked by numerous chromosomal alterations involving several key genes. Notably, the tumor harbored a homozygous deletion of the CDKN2A locus, a well-established oncogenic event in melanoma pathogenesis.²² Amplification of RICTOR on 5p13.1, also identified in our case, occurs in a subset of melanomas and has been associated with poor prognosis and the development of resistance to certain therapeutic agents.^23,24

Interestingly, the tumor also demonstrated heterozygous loss of the EXT2 locus on chromosome 11p11.2. ETX2, and its homolog EXT1, plays a pivotal role in the biosynthesis of heparin sulfate, a vital component of the extracellular matrix and an important regulator of cartilage production.²⁵ EXT2 and EXT1 have been implicated in hereditary multiple osteochondromas (also known as hereditary multiple exostosis), as well as secondary, and rarely primary, chondrosarcomas.^26–29 To our knowledge, the exact 12.193 Mb deletion observed in our case, encompassing chromosome 11p12-p11.2, has not been reported in osteochondromas and chondrosarcomas. However, deletions involving EXT2, including similarly large deletions that involve surrounding genes, have been reported in individuals with Hereditary Multiple Osteochondromas.^30–33 Disruption of EXT2 in these individuals can occur through a variety of mechanisms, including non-sense mutations, missense variants, frameshifts, splice site mutations, and deletions, and are thought to lead to loss-of-function of EXT2. There is sufficient evidence that EXT2 acts in a dosage-sensitive manner, such that loss of a single copy of the gene, as observed in our case, can be pathogenic.^31,32,34 Targeted functional studies would be necessary to elucidate the role, if any, of EXT2 in cartilaginous differentiation in melanoma.

Cartilaginous and osteocartilaginous differentiation in melanoma poses diagnostic challenges owing to its resemblance to other primary tumors. Potential histopathological mimics includes the spectrum of primary cartilaginous or osteocartilaginous lesions, including chondrosarcoma, chondroma, osteochondroma, subungual exostosis, bizarre parosteal osteochondromatous proliferation, and synovial chondromatosis. Generally, these lesions can be distinguished on the basis of their primary origin in the bone or joint space. These melanomas could also show overlapping features with calcified chondroid mesenchymal neoplasms, a recently described soft tissue lesion that frequently involves the hands and digits and has a prominent chondroid matrix. In contrast to melanoma, these tumors typically have distinctive calcifications, often contain giant cells, and are characterized by frequent FN1 fusions.^35,36 Identification of a nested population of tumor cells and the presence of an overlying in situ component are key morphologic clues to a diagnosis of melanoma. Immunohistochemistry for neuromelanocytic markers can be helpful to confirm the diagnosis, although some tumors may lose expression of Melan-A and HMB-45, as noted in our case and in other studies.^12,21 The cartilaginous component of our case exhibited strong and diffuse expression of PRAME. The utility of PRAME in differentiating melanoma with cartilaginous differentiation from a primary cartilage tumor, however, remains unclear. This is primarily due to the scarcity of studies examining PRAME immunohistochemical expression in chondrosarcoma and other cartilage tumors. Despite this, findings from two studies suggest a low level of PRAME mRNA expression in chondrosarcoma tumors and cell lines, indicating that PRAME may be a helpful marker in this distinction.^37,38

In conclusion, this case highlights a rare morphological variant of melanoma and offers insight into its molecular genetics. Awareness of this phenomenon is critical for acute diagnosis and appropriate clinical management. Additional molecular studies could help elucidate the underlying biological drivers of this distinctive histopathological presentation and could reveal new therapeutic targets for these uncommon melanomas.