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Introduction

Melanoma is a highly malignant tumor that originates from melanocytes and can occur in different sites or tissues, including the skin, ocular uvea, pia mater and mucosa, for example, that of the digestive, respiratory and genitourinary tracts. Although malignant melanoma (MM) accounts for only 1% of all diagnosed cutaneous cancers (1), it is responsible for the highest number of skin cancer-associated deaths. MM of the external ear is rare. Among head and neck MMs, the majority (70–90%) occur on the face, mainly on the cheek, while the external ear accounts for only 7% of cases (2). The incidence of primary melanoma of the external ear is increasing, as is the incidence of cutaneous melanoma (3); however, case reports of primary external ear MM are rare.

MM primarily metastasizes through lymphatic and hematologic routes, and distant metastases are most commonly found in the lungs, liver and brain. Gastrointestinal metastases of MM are rare, and as the main site of gastrointestinal metastasis is the small intestine, metastasis of MM to the stomach is even rarer (4). Due to its rarity, to the best of our knowledge there are no case reports or series evaluating the gastric metastasis of MM from the external ear that provide useful disease characterization to help clinicians choose appropriate treatments.

The present study reports the case of a patient with MM originating from the left auricle with initial metastasis to the stomach and descending duodenum, followed by multiple metastases to the brain, lungs, peritoneum, liver and left adrenal gland. The molecular genetic alterations of the tumor were investigated to help select the appropriate targeted therapeutic regimen, and the presence of multiple mutated genes was identified.

Case report

Materials and methods

BRAF V600E mutation was detected using a Human BRAF V600E Mutation Detection Kit (Production lot number 11221112601X; Amoy Diagnostics Co., Ltd.) for medical diagnosis. According to the manufacturer's instructions, amplification of the reference gene (internal control) and BRAF V600E mutation in samples were detected by amplification-refractory mutation system polymerase chain reaction (ARMS-PCR) using HEX and FAM fluorescent channels, respectively. The primers used for PCR were provided as part of the kit. To achieve reliable results, the HEX signal of the sample to be tested should be a positive curve, with a Cq value of between 13 and 21. The cutoff value of Cq for BRAF V600E mutation is 30. Immunohistochemical staining was performed on 4 µm-thick paraffin sections. The ready-to-use primary antibodies (MAB-0098, MAB-0275, Kit-0007, MAB-0671, Kit-0024, RMA-0815, MAB-0743, MAB-0742, MAB-0707, MAB-0672, MAB-0735, MAB-0716 and MAB-0717; MXB Biotechnologies Co., Ltd.) and an EliVision plus Detection Kit (KIT-9901; MXB Biotechnologies Co., Ltd.) were used for immunohistochemical staining.

Case presentation

A 58-year-old female patient was hospitalized at the People's Hospital of Xiangyun County (Xiangyun, China) in March 2019 due to a black mass the approximate size of a fava bean behind the left ear that had been present for ~5 years with itching but no tinnitus, stuffiness, hearing loss or pus discharge from the external ear canal. Upon physical examination, it was observed that the black mass was soft with clear boundaries and an absence of pain and redness, and both auricles exhibited normal morphology. Five days after initial hospitalization, mass resection and arbitrary flap plasty were performed. The resected mass comprised hard grayish-black tissue, 1.5×1×0.8 cm3 in size, with a black protrusion of diameter 0.8 cm in the center (Fig. 1A). The mass was diagnosed as nodular MM of the left auricle by intraoperative frozen section examination. When observed under a microscope, nestlike, nodular and diffuse melanocyte dysplasia was observed, with tumor cells displaying marked pleomorphism and high proliferative activity (Figs. 1B and C). Immunohistochemical staining showed that the tumor cells were positive for human melanoma black 45 (HMB45; Fig. 1D), melan-A (Fig. 1E) and S-100 (Fig. 1F) but negative for pan cytokeratin (CK-pan), leukocyte common antigen and p40. Formalin-fixed paraffin-embedded tissue sections were further analyzed to detect genetic variations. Fluorescence in situ hybridization (FISH) revealed MYB deletion (Fig. 2A), p16 deletion (Fig. 2B), MYC amplification (Fig. 2C), PTEN deletion (Fig. 2C) and no marked changes in the copy numbers of Ras responsive element binding protein 1 (RREB1; Fig. 2A) and cyclin D1 (CCND1; Fig. 2B). ARMS-PCR revealed the BRAF V600E mutation (Fig. 3), and no common NRAS (G12D, G12S, G13D, G13R, G12C, G12V, G12A, G13V, A59D, Q61R, Q61K, Q61L, Q61H, K117N, and A146T) mutations were found. Furthermore, DNA sequencing revealed no mutations in exons 9, 11, 13, or 17 of c-KIT. PCR followed by high-resolution melting analysis showed that the tumor was negative for seven microsatellite instability biomarkers, namely activin A receptor type 2A; BTB domain containing 7; death inducer-obliterator 1; MRE11 homolog, double strand break repair nuclease; ryanodine receptor 3; SEC31 homolog A, COPII coat complex component; and sulfatase 2. These findings indicated microsatellite stability.

Figure 1. Pathological evaluation of MM of the external ear in the present case. (A) Resected specimen of the left auricle. Hematoxylin and eosin staining of the resected auricular MM specimen at (B) ×200 and (C) ×400 magnification. Immunohistochemical staining showed that tumor cells in the MM were positive for (D) human melanoma black 45, (E) melan-A and (F) S-100; magnification, ×400. MM, malignant melanoma.

Figure 2. Copy number alterations of multiple genes in the auricular MM detected by fluorescence in situ hybridization. The MM was found to be (A) positive for 6q23 (MYB) deletion (44%) and negative for 6p25 (RREB1) amplification (4%), (B) positive for 9p21 (P16) deletion (80%) and negative for 11q23 (CCND1) amplification (2%) and (C) positive for 8q24 (MYC) amplification (average 2.8 signals) and 10q23 (PTEN) deletion (70%) (×1,000 magnification). The specific validated cutoff values for the probes used in the assay are as follows: 1 red signal (red arrow) in ≥40% of nuclei is considered an abnormal loss of MYB; >2 green signals (green arrow) in ≥29.0% of nuclei is considered an abnormal gain of RREB1; ≤1 red signal (red arrow) in ≥20% of nuclei is considered an abnormal loss of P16; >2 green signals (green arrow) in ≥38.0% of nuclei is considered an abnormal gain of CCND1; average >2.5 green signals (green arrow) in nuclei is considered an abnormal gain of MYC; 1 red signal (red arrow) in ≥20% of nuclei is considered an abnormal loss of PTEN. MM, malignant melanoma; RREB1, Ras responsive element binding protein 1; CCND1, cyclin D1.

Figure 3. Amplification curve of BRAF V600E mutation in the auriclar MM detected by ARMS-PCR. Amplification of the reference gene (internal control) and BRAF V600E mutation were detected by ARMS-PCR using HEX and FAM fluorescent channels, respectively. The Cq values for the reference gene and BRAF V600E were 16.5 and 22.8, respectively, suggesting that the present case of MM was positive for the BRAF V600E mutation. MM, malignant melanoma; ARMS-PCR, amplification-refractory mutation system polymerase chain reaction.

The patient was hospitalized again at the People's Hospital of Xiangyun County in March 2020 due to epigastric pain, nausea, vomiting and abdominal distension. Protruding lesions with erosion and melanin deposition were observed in the gastric body, gastric antrum and descending part of the duodenum by gastroscopic examination (Fig. 4). Pathological biopsy revealed gastric mucosa hyperemia, nestlike cell clusters in the lamina propria, and lymphocyte and plasma cell infiltration (Fig. 5A-C). Helicobacter pylori was not detected. Immunohistochemical staining showed that the lesion cells were positive for HMB45 (Fig. 5D), melan-A (Fig. 5E) and S-100 (Fig. 5F), weakly positive for CD56 (Fig. 5G), and negative for CK-pan, Syn and CgA. The Ki-67 proliferation index was ~40% (Fig. 5H). Due to the medical history, histopathologic features and expression of protein markers in the biopsy specimen, the patient was diagnosed with metastatic external ear MM. Subsequently, the patient received targeted treatment with vemurafenib, which was administered during multiple immunotherapy sessions at a specialized oncology hospital.

Figure 4. Gastroscopic findings. Black protruding lesions in the (A) gastric body and (B) antrum.

Figure 5. Pathological evaluation of the gastric metastatic malignant melanoma. Hematoxylin and eosin staining of the biopsy specimen of the gastric mucosa at (A) ×100, (B) ×200 and (C) ×400 magnification. Immunohistochemical staining showed tumor cells positive for (D) human melanoma black 45, (E) melan-A, (F) S-100, (G) CD56 and (H) Ki-67; magnification, ×400.

The patient experienced abdominal swelling for ~2 weeks in February 2021. Computed tomography examination revealed multiple hyperdense shadows in the brain (Fig. 6A), bilateral small pulmonary nodules in the lungs (Fig. 6B), diffuse slightly hyperdense nodules of varying sizes in the liver (Fig. 6C), space-occupying lesions in the left adrenal gland (Fig. 6D) and peritoneal thickening (Fig. 6E), suggesting systemic metastases of the tumor. The patient had seroperitoneum in which malignant tumor cells were detected by cytological examination (Fig. 7A). Immunohistochemical staining showed that the tumor cells were positive for HMB45 (Fig. 7B), melan-A (Fig. 7C), S-100 (Fig. 7D), Ki-67 (50%) (Fig. 7E) and vimentin (Fig. 7F), and the peritoneal mesothelial cells were positive for CK-pan (Fig. 7G), calretinin (Fig. 7H) and mesothelial cell (Fig. 7I), indicating metastatic MM.

Figure 6. Computed tomography scans of the patient. Metastatic lesions in the (A) brain, (B) lungs, (C) liver, (D) left adrenal gland and (E) peritoneum were detected.

Figure 7. Pathological evaluation of the metastatic malignant melanoma cells in the seroperitoneum. (A) Hematoxylin and eosin staining of a paraffin-embedded cell block of the seroperitoneum (×400 magnification). Immunohistochemical staining showed tumor cells positive for (B) human melanoma black 45, (C) melan-A, (D) S-100, (E) Ki-67 and (F) vimentin and peritoneal mesothelial cells positive for (G) pan cytokeratin, (H) calretinin and (I) mesothelial cell (×400 magnification).

In March 2021, the patient developed secondary epilepsy, hypoproteinemia, electrolyte disturbance and cancerous cachexia. The patient abandoned treatment and died two days after discharge from the hospital (Fig. 8).

Figure 8. Timeline of the history of the present case. CT, computed tomography.

Literature review

A total of 6,577 cases of MM of the external ear reported in 11 articles were reviewed in the present study (Table I) (3,5–14). The cases in all articles had MM of the external ear, and the specific site was reported in some of the articles. The tumors primarily occurred in the helix. Analysis of these cases indicated that MM of the external ear frequently occurs in middle-aged and older adults and occasionally in adolescents. Men (5,603 cases) were more frequently affected than women (974 cases). Patients with MM of the external ear were mostly asymptomatic or had some local symptoms at diagnosis, including ear swelling, otorrhea, hearing loss and bleeding. Regional lymph node metastasis was common, and the other metastatic sites were primarily the liver, lung, brain, parotid gland and retromandibular tissue. Regarding prognosis, a long disease duration and high tumor stage were unfavorable factors for survival.

Table I. Cases of malignant melanoma of the external ear reported in the literature (n=6,577).

Table I.

Cases of malignant melanoma of the external ear reported in the literature (n=6,577).

First author/s, year	Cases, n	Age, years	Sex	Primary site	Clinical symptoms	Metastatic sites	Outcomes	(Refs.)
Amando García et al, 2003	1	59	Male	External ear canal	Earache, suppurative otorrhea, preauricular swelling	Parotid gland	Disease-free survival for 9 months	(5)
Hannan and Parikh, 2006	1	40	Male	External ear canal	Deafness	Retromandi bular tissue	N/A	(6)
Gowthami et al, 2014	1	11	Female	Right post-auricular	Pain, swelling, intermittent bleeding	Right temporal bone	Died	(7)
Toia et al, 2015	845	59.4 (mean)	Male (78%), female (22%)	Helix (57%); earlobe (17%); post-auricular (9%); auricle (6%); antihelix and scapha (4%); external ear canal (0.8%); other (6.2%)	N/A	Regional lymph nodes (10%)	Five-year survival rate: stage 0, 100%; stage I, 71%; stage II, 53%; stage III, 0%	(8)
Frost et al, 2017	45	63 (median)	Male (80%), female (20%)	External ear	N/A	Sentinel lymph nodes (7%); distant metastasis (4.4%)	Survival rate: Overall, 80%; stage I/II,84%; stage III/ IV, 56%	(9)
Deep et al, 2017	5,481	66.7 (mean)	Male (86.5%) female (13.5%)	External ear	N/A	N/A	Survival rate: Overall, 90%; stage I, 95.3%; stage II, 81.1%; stage III, 56.6%; stage IV, 20.5%	(3)
Harrison et al, 2017	41	61 (median)	Male (73%), female (27%)	External ear	N/A	Liver and brain (5%)	Survival rate: Overall, 90%; 5-year, 100%; 10-year, 71%	(10)
Straccia, et al 2020	1	43	Female	Right auricle	N/A	Pleura	N/A	(11)
Truong et al, 2020	156	62.5 (median)	Male (86%), female (14%)	Helix (61.5%); earlobe, (13.5%); other (25%)	N/A	Sentinel lymph nodes (14.1%)	Survival rate, 74.4%	(12)
Fiorio et al, 2021	4	69 (mean), 67.5 (median)	Male (25%), female (75%)	Right earlobe (50%); left helix (50%)	Asymptomatic	Regional lymph nodes (25%)	During the follow-up, with no signs of disease progression	(13)
Pająk et al, 2022	1	43	Female	Left helix	Asymptomatic	N/A	Lost to follow-up	(14)

[i] N/A, no relevant information available.

A detailed literature review of 22 cases of gastric metastatic melanoma was performed (Table II) (15–36). The sex of one patient was not reported, and of the remaining patients, 14 (66.7%) were male and 7 (33.3%) were female. The mean age at diagnosis was 63.1 years (range, 36–89 years). Excluding five patients with unclear metastatic site in the stomach, the majority of the gastric metastatic melanomas occurred in the gastric body (41.2%) and fundus (29.4%), and the rest occurred in the gastric antrum, cardia and pylorus. The whole stomach was involved in one case. On the basis of the articles containing clear reports of 19 patients' symptoms, it was found that the main gastrointestinal symptoms were black stools or hematemesis (52.6%), abdominal pain (36.8%) and nausea or emesis (26.3%). Less common symptoms included anorexia and abdominal distention. Endoscopic or intraoperative findings were mostly masses, nodules, protruding lumps or polyps (77.3%), ulcerations (45.5%) and melanin pigmentation (31.8%). Multiple mucosal erosions were less common manifestations. The primary site was not reported for one patient, and the skin was the most common primary site in the remaining patients (16 cases, 72.7%), including eight with acral MM. Two cases of gastric metastatic MM originated in the eye, two in the mucosa and one in the sphenoid sinus. Following the exclusion of six patients with missing time-to-metastasis data, the mean and median time intervals to metastasis were 4.4 and 2 years, respectively, with a range from 0.5 to 15 years. Individual differences in tumor metastasis among the patients are evident.

Table II. Cases of gastric metastatic malignant melanoma reported in the literature (n=22).

Table II.

Cases of gastric metastatic malignant melanoma reported in the literature (n=22).

First author/s, year	Sex	Age, years	Metastatic site	Clinical symptoms	Endoscopic or intraoperative findings	Primary site	Time interval between primary and metastatic lesions	(Refs.)
Davis and Zollinger, 1960	Female	76	Greater gastric curvature	Anorexia, nausea and black stools	Multiple polypoid lesions	Upper arm	1 year	(15)
Morini et al, 1980	Female	77	Posterior wall of the gastric body	Abdominal pain, dyspepsia, lack of appetite, and black stools	Protruding lump	Left hip	6 years	(16)
Mimica and Tomić, 2002	Male	51	Gastric body and antrum	Abdominal pain	Multiple polypoid lesions	N/A	N/A	(17)
Malladi et al, 2005	Male	80	Fundus, upper part and pylorus of the stomach	Abdominal pain, dyspepsia and weight loss	Multiple black nodules with ulceration	Sole of the right foot	2 years	(18)
Matsubayashi et al, 2005	Male	53	Upper part of the stomach	N/A	Dark brown lesions with clear boundaries	Gums	Metastatic at diagnosis	(19)
Wu et al, 2011	Male	72	Stomach	Dysphagia	Superficial ulcers and ulcerative nodules	Esophagus	Metastatic at diagnosis	(20)
Law et al, 2011	Female	84	Stomach	Hematemesis	Multiple pigmented nodules and mucosal swelling	Right foot	1 year	(21)
Eivazi-Ziaei and Esmaili, 2014	Male	56	Lesser gastric curvature	Abdominal	Ulcers and lumps pain	Right heel	2 years	(22)
El-Sourani et al, 2014	Female	43	Lesser gastric curvature	Black stools and anemia	Lump	Right breast	2 years	(23)
Zhao et al, 2015	Male	57	Stomach	Diarrhea	Multiple erosion and lumps	Sphenoid sinus	Half a year	(24)
Ruiz-Cuesta et al, 2014	N/A	49	Gastric body	Epigastric pain, nausea, weight loss	Ulcerated lump	Right lower limb	13 years	(25)
Morita et al, 2019	Male	55	Whole stomach	N/A	Mucosa covered by tumor cells with pitting and melanin pigmentation	Uvea	14 years	(26)
Farshad et al, 2018	Male	89	Gastric cardia	Hematochezia	Bleeding mass and ulcerations	Chest	15 years	(27)
Santos-Seoane et al, 2019	Male	79	Gastric fundus	N/A	Polypoid lesion	Left forearm	3 years	(28)
Sachdeva et al, 2020	Male	59	Gastric body, fundus and antrum	Black stools and weight loss	Multiple black ulcerated nodules	Multiple skin areas throughout the body	10 months	(29)
Anandabaskaran et al, 2020	Male	68	Gastric incisura	Abdominal pain and hematochezia	Pitted ulcerations with melanin pigmentation around the periphery	Right foot	7 years	(30)
Groudan et al, 2020	Female	66	Gastric antrum	Nausea and hematemesis	Ulcerations and fungoid mass	Vulva	Metastatic at diagnosis	(31)
Yoshimoto et al, 2021	Female	82	Gastric body	Black stools	White semicircular ulcerated mass	Left toe	N/A	(32)
Monti et al, 2021	Male	56	Stomach	Severe dyspepsia	Ulcerations and plaques	Glans penis	Metastatic at diagnosis	(33)
Zhu et al, 2022	Male	36	Gastric body and fundus	Anorexia, nausea and emesis	Hemispherical protrusive lesion	Sole of the right foot	6 months	(34)
Maksimaityte et al, 2022	Male	46	Gastric body and fundus	Nausea and abdominal enlargement	Multiple red polyps	Right ear auricle	1 year	(35)
Bharwad et al, 2022	Female	55	Stomach	Abdominal pain, hematochezia, dizziness and fatigue	Multiple small pigmented lesions	Choroid of the right eye	2 years	(36)

[i] N/A, no relevant information available.

Discussion

Primary MM of the external ear is a rare type of melanoma, accounting for ~1% of cutaneous MM and 7% of MM of the head and neck (2). In the present review it was identified that tumor lesions mainly occurred in the helix and anthelix, followed by the earlobe (lobules auriculae) and auricle (concha auriculae). The external auditory canal was a less common site. The most common subtype of external ear MM was superficial spreading melanoma (40.1%), followed by malignant freckle-like nevus (33.7%) and nodular melanoma (16.4%) (8,37). MM of the external ear is primarily observed in the white population (99.3%), with a much lower incidence (<0.5%) in the non-white population (37). Melanin pigmentation is well known for protecting the skin from the harmful effects of UV radiation. A negative association exists between the degree of skin pigmentation and the incidence of external ear MM (37). The average incidence rates of MM are 1.015 cases per 100,000 men and 0.126 cases per 100,000 women (37). The patient described in the present case report is an Asian woman, and MM is extremely rare in individuals with these demographic characteristics.

Patel et al (37) reported that the 5- and 10-year disease-specific survival rates for MM of the external ear are 89.4 and 84.8%, respectively, and another study suggested that the 5-year overall and cancer-specific survival rates for MM of the outer ear are 78.8 and 90.0%, respectively (3). Patients with a malignant freckle-like nevus have been shown to have favorable survival outcomes, while those with nodular melanoma generally have generally poor outcomes (37). It has been reported that the time of cancer-specific death can be independently predicted by age, Breslow thickness, stage, ulceration and lymphatic and distant metastasis (3). Certain studies have implied that MM of the external ear may be more aggressive than other head and neck MMs (7,38), while others contend that anatomical location does not affect tumor recurrence rates and survival outcomes (3,37,39).

MM is a highly aggressive cancer that metastasizes relatively early, particularly to the lymph nodes, brain, liver and lungs. The preauricular and postauricular lymph nodes, anterior and posterior regions of the neck and parotid gland are common sites of local metastasis for melanoma of the external ear. Consistent with the systemic review of external ear MM published by Toia et al (8), the literature review in the present study observed that regional lymph nodes were the most frequent sites of metastasis, and distant metastatic sites were primarily liver, lung, brain and other head and neck tissues. However, gastrointestinal metastases of MM are uncommon (4,40,41). As reported by López et al (42), gastrointestinal metastases of MM involve the small intestine (51–71%), stomach (27%) and colon (22%). In the present case and literature review, it was found that gastric metastatic MM typically occurs in the gastric body and fundus, with various manifestations observed in endoscopy or surgery, including masses, nodules, bulges, ulcerations, polyps, mucosal pigmentation and erosions. A review of studies on small intestinal melanomas suggested that 60% of deceased patients with MM had gastrointestinal metastases, but only 1.5–4.0% were detected before death (43). The present retrospective literature study revealed that a total of 22 cases of gastric MM were reported between 1960 and 2022, including only one patient with a primary lesion in the ear, which was a 46-year-old white male. To the best of our knowledge, this is the first case report of gastric metastasis from auricular MM in a non-white female.

Prophylactic removal of nevus of the external ear is advocated in the literature to prevent the development of MM (44). Approximately 8% of cases of MM in the ear are non-pigmented, which can easily lead to misdiagnosis. As these tumors are insensitive to radiotherapy, wide local excision should be performed as soon as possible if malignancy is suspected. Surgery, with or without lymph node dissection, remains the preferred treatment strategy for MM (8). Numerous adjuvant therapies have been investigated for the treatment of cutaneous MM following complete surgical excision. The US Food and Drug Administration (FDA) has approved ipilimumab, nivolumab and pembrolizumab for the adjuvant treatment of MM at stage III and above (45). Immune checkpoint inhibitors represent a promising new type of systemic therapy for the adjuvant treatment of MM. Regarding the surgical treatment of MM from the external ear, the safe margin of resection remains unclear. A minimum resection margin of 1 cm is commonly reported; however, some authors advocate a resection margin of 2 cm if the tumor thickness is >2.0 mm. There is a trend towards surgical approaches with reduced resection margins, with the advised minimum extent being >0.5 cm (3,46). The resection margin in the present case was much smaller than the minimum resection standard advocated in the aforementioned studies, which may a primary reason for the postoperative metastasis of the tumor.

Positive results for specific immunohistochemical markers, including HMB45, melan-A, S-100, SRY-box transcription factor 10, melanoma-associated antigen (PNL-2), tyrosinase and melanocyte inducing transcription factor, are critical for the diagnosis and differential diagnosis of MM (47). HMB45 is a glycoprotein expressed in melanosomes with extremely high specificity; melan-A, PNL-2 and tyrosinase also have high specificity; while S-100 is more sensitive but less specific for the detection of MM. The appropriate marker for detection and diagnosis can be selected on the basis of the marker characteristics. In the present study, HMB45, melan-A, and S-100 were found to be expressed in the tumor cells in the primary lesion, gastric metastatic lesion and seroperitoneum of the patient.

The initiation and development of MM involve multiple genes. BRAF gene mutation is the most prevalent genetic alteration in 40–60% of cutaneous MM cases, and this mutation results in constitutive activation of the MAPK pathway (48–50). Vemurafenib, dabrafenib and encorafenib are US FDA-approved inhibitors targeting the BRAF V600 mutation, and a combination of BRAF inhibitors and MEK inhibitors, including trametinib and cobimetinib, has become the standard treatment for advanced MM with BRAF V600 mutation (51,52). Furthermore, atezolizumab plus vemurafenib plus cobimetinib was approved by the FDA for patients with BRAF V600 mutation-positive unresectable or metastatic MM in July 2020 (53). Therefore, molecular testing for BRAF mutations in MM is critical for selecting the most appropriate treatment. Genetic information is rarely mentioned in reported cases of MM of the external ear, and the present case was positive for the BRAF V600E gene mutation. BRAF mutations are associated with a higher tumor stage, lymph node metastasis and an increased depth of tumor infiltration (54). Brain metastases are more likely to occur in patients in which the MM has BRAF mutations than in those with wild-type BRAF (55). Consistent with this, 11 months following the gastric metastasis of external ear MM, the involvement of multiple organs, including the brain, had occurred in the present case.

Numerous molecular genetic studies have demonstrated that although BRAF mutations have been detected in both MM and melanocytic nevi, the genetic alterations of the two presentations differ substantially. MM frequently shows a large number and variety of chromosomal abnormalities, including 6q23 (MYB), 6q25 (RREB1), 9p21 (p16), 11q23 (CCND1), 8q24 (MYC) and 10q23 (PTEN), which can be amplified, deleted or translocated (56). By contrast, these genetic loci are rarely changed in melanocytic nevi. Multiple molecular testing by FISH confirmed the amplification of the MYC gene and deletion of the MYB, p16 and PTEN genes in the present case. Additionally, RREB1 and CCND1 copy numbers were not markedly changed.

The proto-oncogene c-MYC is strongly associated with various types of cancer. It has been demonstrated that the copy number of c-MYC is increased in the tumor region of nodular MM (57). In a study of 43 cases of uveal melanoma, extra copies of c-MYC were detected in 70% of cases (58). Of the 30 patients with extra copies of c-MYC, 13 (43%) had amplification, 14 (47%) had an intermediate relative increase in copy number, and 3 (10%) had a simple gain of chromosome 8 (58). In a study of 44 cases of primary acral melanoma, c-MYC was found to be amplified in 54.5% of cases (59). In addition, c-MYC gene expression may be positively associated with the metastatic potential of MM (57). An association has also been identified between larger tumor size and c-MYC amplification (58).

The MYB gene is localized in chromosome 6q22-q23. Deletions and translocations involving this region of the long arm of chromosome 6 occur frequently in MM, and rearrangements of the MYB gene have been reported in MM (60). Romano et al (61) observed deletion of the MYB gene in five of seven cases (71%) of melanoma of the nail apparatus, with the percentage of abnormal cells ranging from 52 to 94%. Two of the patients in the study had lymph node metastasis and died from widespread metastatic disease, and these two patients had amplification of RREB1, CCND1 and MYC as well as MYB deletion (61).

As a tumor suppressor, p16 has been shown to play a crucial role in tumor progression rather than initiation. In healthy cells, p16 blocks the phosphorylation of retinoblastoma protein by binding to CDK4/6, leading to cell cycle arrest. Dysfunction of the p16 protein diminishes its ability to regulate cell proliferation, resulting in uncontrolled cell division and the growth and development of tumors (62). p16 alterations frequently occur in sporadic melanomas (62). In a study of vertical growth phase melanoma, absent or minimal nuclear p16 protein expression was found in 45% of primary melanomas, and further loss of p16 expression was observed in 77% of metastatic lesions; the absence of p16 expression independently predicted poor survival in the patients (62).

PTEN is a well-known tumor suppressor that is frequently mutated, inactivated or deleted in various malignancies. The MEK/ERK pathway contributes to the inhibition of PTEN by activating c-Jun, which activates the PI3K/AKT pathway, causing apoptosis resistance, tumor growth and metastasis in melanoma cells (63,64). Loss of PTEN function has been reported in 35% of melanomas and is often detected in advanced melanomas along with BRAF mutations (65). Dankort et al (66) induced the expression of BRAF V600E specifically in the melanocytes of mice; the mice developed benign melanocytic hyperplasias that did not progress to melanoma within 15–20 months. By contrast, when the expression of BRAF V600E was combined with PTEN gene silencing, melanoma with 100% penetrance, a short latency period and metastases to the lymph nodes and lungs developed, suggesting that the BRAF V600E mutation combined with PTEN deletion induces MM metastasis (66). It has been reported that PTEN deletion is a possible predicting factor of intrinsic resistance to BRAF inhibitor therapy (67). Notably, BRAF inhibitor treatment was ineffective in the present patient. In addition, PTEN deletion has been described as a driver of immune evasion, with PTEN deletion in MM being associated with the increased expression of programmed death-ligand 1 and immunosuppressive cytokines, decreased T-cell infiltration, and resistance to T-cell-mediated tumor cell lysis (64). PTEN deletion has also been suggested to mediate immune evasion by the attenuation of tumor antigen cross-presentation, resulting in T-cell exclusion and indirectly influencing B-cell function (67).

In conclusion, the present report describes a rare case of MM in an Asian female with postoperative metastasis to the stomach and descending duodenum from the auricle and secondary metastases to multiple organs, including the brain, lungs, liver, left adrenal gland and peritoneum. Molecular testing revealed genetic aberrations in BRAF, MYB, p16, MYC and PTEN. The small resection margin, nodular histological type and number of genetic aberrations may be strongly associated with high-metastatic potential, resistance to targeted therapy and poor prognosis in this case.

Acknowledgements

The authors would like to thank Guangzhou LBP Medical Technology Co., Ltd. (Guangzhou, Guangdong, China), for providing FISH technical services.

Funding

The present study was supported by the National Natural Science Foundation of China (grant no. 82160582), the Yunnan Province Fundamental Research Project (grant no. 202201AT070003) and the Scientific Research Foundation of Dali (grant no. 2021KBG036).

Availability of data and materials

All data generated or analyzed during this study are included in this published article.

Authors' contributions

YCY detected the gene mutation, collected the pathological images, drafted the manuscript and analyzed the data for the literature review. YL and YP contributed to the pathological diagnosis and clinical data collection. BG contributed to the analysis and interpretation of the case report and revised the manuscript. YCY, YL, YP and BG confirm the authenticity of all the raw data. All authors read and approved the final version of the manuscript.

Ethics approval and consent to participate

The present study was approved by the Ethics Committee of the First Affiliated Hospital of Dali University (approval no. 20221202001). Informed consent was obtained from the patient's family.

Patient consent for publication

Consent for publication was obtained from the patient's family.

Competing interests

The authors declare that they have no competing interests.

References