Primary cutaneous NUT carcinoma with BRD4::NUTM1 fusion

Nuclear protein in testis (NUT) carcinoma, molecularly defined by the NUTM1 gene rearrangement, is most commonly reported in young adults in the sinonasal tract, nasopharynx, or thorax. At these sites, NUT carcinoma is an extremely aggressive malignancy with dismal prognosis. Recently, five cases of primary cutaneous NUT adnexal carcinoma have been reported with BRD3 and NSD3 fusion partners. Although NUT adnexal carcinomas are shown to have metastatic potential, they may behave less aggressively than extracutaneous NUT carcinomas. We report a case of a 59‐year‐old man who underwent a biopsy of a 3‐cm plantar mass, which showed BRD4::NUTM1 fusion. The tumor was a poorly differentiated dermal neoplasm showing cytologic atypia, large vesicular nuclei with prominent nucleoli, conspicuous mitotic activity, and foci of necrosis. Immunohistochemically, the tumor showed positivity for keratins, EMA, SOX10, and NUT, with patchy smooth muscle actin. Molecular testing revealed BRD4::NUTM1 rearrangement. With no alternative primary identified by imaging, a diagnosis of primary cutaneous NUT carcinoma was favored. We hope to contribute to the limited body of knowledge on this entity, with emphasis on recognition as well as studying and defining its prognostic differences from extracutaneous NUT carcinomas.

59-year-old man who underwent a biopsy of a 3-cm plantar mass, which showed BRD4::NUTM1 fusion.The tumor was a poorly differentiated dermal neoplasm showing cytologic atypia, large vesicular nuclei with prominent nucleoli, conspicuous mitotic activity, and foci of necrosis.Immunohistochemically, the tumor showed positivity for keratins, EMA, SOX10, and NUT, with patchy smooth muscle actin.Molecular testing revealed BRD4::NUTM1 rearrangement.With no alternative primary identified by imaging, a diagnosis of primary cutaneous NUT carcinoma was favored.
We hope to contribute to the limited body of knowledge on this entity, with emphasis on recognition as well as studying and defining its prognostic differences from extracutaneous NUT carcinomas.

| INTRODUCTION
Nuclear protein in testis (NUT) carcinoma represent a unique subset of epithelial malignancies defined by rearrangement involving the NUT gene.2][3] These tumors have been reported predominantly in midline structures such as the sinonasal tract, nasopharynx, and thorax.They typically affect young adults and show an overall dismal prognosis.
The characteristic histopathology of NUT carcinoma is the proliferation of sheets and nests of small to intermediate sized cells with undifferentiated "small round cell" morphology with abrupt keratinization.
5][6] Even though primary cutaneous NUT A poster of these findings was presented at the American Society of Dermatopathology 60th Annual Meeting on October 7, 2023.carcinomas have demonstrated metastatic potential, their clinical behavior is so far reported to be indolent in contrast to their extracutaneous counterparts.This case report adds to the growing literature by presenting primary cutaneous NUT carcinoma with BRD4::NUTM1 rearrangement, displaying a high-grade undifferentiated morphology.It remains to be seen whether this morphology and/or fusion will correlate with a more aggressive clinical course.Unlike their extracutaneous counterparts, the prognostic implications of fusion partners in cutaneous NUT carcinomas remain unknown.

| CASE REPORT
A 59-year-old White man with no prior relevant dermatologic history presented with a "bump" on the first metatarsophalangeal (MTP) joint, which was clinically diagnosed as a verruca and subsequently failed multiple treatments with cryotherapy.A year later, the patient presented with a large fungating tumor on the right plantar foot.On MRI, a 3.0 Â 2.5 Â 2.9 cm avidly enhancing mass was located plantar to the first MTP joint involving the skin and subcutaneous tissue but not involving the adjacent tendons or bones (Figure 1).CT imaging of the chest, abdomen, and pelvis was negative for metastases or alternative primary tumor.At this point, the patient underwent a biopsy for pathologic evaluation.
Histologic sections revealed a dermal-based, cellular, poorly differentiated malignant neoplasm forming infiltrative trabeculae and nodules within a desmoplastic stroma (Figure 2A).intact INI-1 expression, and negativity for desmin, CD34, and S100 (not shown).However, BRD4::NUTM1 rearrangement was identified by next-generation sequencing (ArcherDX, Inc.), and further immunohistochemistry showed positive NUT staining (Figure 3).With no alterative primary identified by imaging, and considering that SOX10 and cytokeratin co-immunoreactivity was previously reported in NUT carcinoma, 6 a diagnosis of primary cutaneous NUT carcinoma was favored.Current case Age, sex, site 7, female, shoulder 46, female, inguinal 4 34, female, inguinal 6 One case not reported 7 Not reported 58, male, foot

Epidermal connection
No connection Not reported 4 One case reported as "dermal-based" 6 One case connected to epidermis Following the biopsy, patient received preoperative chemoradiation with modified Elber protocol, which includes 3 days of IV infusion of low-dose adriamycin followed by radiotherapy of a total tumor dose of 3000 cGy delivered in 10 fractions in an attempt to shrink the tumor prior to resection.
The resection specimen revealed residual tumor cells of similar morphology involving the dermis and extending to the subcutaneous tissue, but with significantly lower mitotic rate (not shown).The resection specimen showed foci of lymphovascular invasion (Figure 2D).
The patient is alive 3 months after preoperative neoadjuvant chemoradiation and 3 months post surgical resection with no evidence of metastatic disease or recurrence.

| DISCUSSION
Primary cutaneous NUT carcinomas are rare; based on our search in PUBMED and EMBASE databases, only five cases have been reported in literature: one with BRD3::NUTM2B fusion, three with BRD3:: NUTM1 gene fusions, and one with NSD3::NUTM1 fusion (Table 1).
Three out of the five cutaneous NUT carcinomas reported thus far have also had lymph node and/or distant metastases (Table 1).In one case of right inguinal cutaneous NUT carcinoma with BRD3::NUTM1 fusion, the patient developed multiple pelvic lymph node metastases and metastases to the left foot followed by multiple metastases on the left leg. 4 Despite lymph node and distant metastases, the patient showed an indolent clinical course of more than 10 years. 4 our case, the patient opted for a conservative resection rather than an amputation.The patient was found to have lymphovascular invasion (Figure 2D), but no lymph node dissection was performed.
However, it is too early to comment on tumor behavioral characteristics such as recurrence or metastatic potential at this stage.
Interestingly, cutaneous NUT carcinomas with BRD3 fusions have been reported to have an adnexal phenotype exemplified by foci of ductal and glandular differentiation abruptly admixed with squamous keratocystic structures reminiscent of follicular infundibular differentiation (Table 1).An important primary cutaneous differential diagnosis to consider is porocarcinoma with YAP1::NUTM1 fusion, which also shows strong NUT nuclear positivity by immunohistochemistry. 8,9 Although foci of poroid differentiation have been reported in cutaneous NUT carcinomas, previous reports have argued that the presence of keratocystic follicular infundibular differentiation defines these tumors as a distinct entity rather than a variant of porocarcinoma. 4,5fferentiating from myoepithelial carcinoma can be challenging, especially in cases with inconspicuous ductal or keratocystic structures, but negative NUT immunostaining as well as detection of EWSR1 or PLAG1 rearrangement favors myoepithelial carcinoma.Furthermore, appropriate immunohistochemical workup should be sufficient to distinguish NUT carcinomas from poorly differentiated squamous cell carcinoma (diffusely p63/p40+, and SOX10 and NUTÀ), epithelioid sarcoma (SOX10, NUT, and INI1À).
Our case of cutaneous NUT carcinoma with BRD4::NUTM1 fusion distinguishes itself from previously reported tumors with BRD3 and survival. 10However, the prognostic significance of fusion partners in primary cutaneous NUT carcinomas is not known.
Based on our molecular analysis, BRD4-NUT protein, which is the product of t (15;19), in our case results from 5 0 cDNA breakpoint in exon 13 of BRD4 gene and 3 0 cDNA breakpoint in exon 3 of the NUTM1 gene (Figure 4).The resultant BRD4-NUTM1 protein contains the bromodomain-like subfamily repeat I and II, bromodomain extraterminal domain, and ZipA C-terminal domain along with the NUTM1 protein (Figure 4).Physiologically, NUTM1 binds with histone acetyltransferase p300, whose function is the acetylation of histone proteins. 11The NUT moiety of the BRD4-NUT fusion protein is shown to interact with p300 leading to transcriptionally inactive hyperacetylated chromatin domains, which results in p53 inactivation. 12BRD4 protein is an epigenetic reader that recognizes acetylated lysine residues in histone proteins and provides a scaffold for other factors, thereby regulating DNA transcription and influencing carcinogenesis. 13,14BRD4 contains two bromodomains that bind transcriptionally active chromatin through associations with histones H3 and H4, 15 and is complexed with BRD3, NSD3, and ZNF through its extraterminal domain. 16nctionally, BRD4-NUT proteins are retained within the nucleus as a result of bromodomain-chromatin interactions. 1[19] BRD4-NUT oncoprotein has been shown to associate with the MYC promoter, thereby maintaining MYC expression in NUT carcinoma. 18 vitro observations in NUT-carcinoma cell lines have shown that siRNA-mediated knockdown of BRD4-NUT or BRD3-NUT protein induces rapid terminal squamous differentiation followed by arrest in proliferation, highlighting its significant role in maintaining an undifferentiated and proliferative state. 1 Because the pathogenesis of NUT carcinoma is primarily due to the effects of the resultant fusion oncoproteins, 11 it can be hypothesized that such fusion-dependant prognostic differences may also be observed in cutaneous NUT carcinomas.However, it is yet to be demonstrated whether BRD4-NUTM1 protein has more robust oncogenic effects than BRD3-NUTM1.Furthermore, it is yet to be seen whether the high-grade cytology would be a consistent feature of these tumors.
In conclusion, this case of primary cutaneous NUT carcinoma with BRD4::NUTM1 fusion adds to our understanding of this rare malignancy, underscoring the importance of continued research into its molecular mechanisms as well as potential fusion-dependent histopathological and prognostic differences.
The tumor was not connected to the epidermis.The tumor cells had scant eosinophilic to clear cytoplasm, large vesicular nuclei with prominent nucleoli, mitotic activity up to six mitoses per 10 high-power fields, and foci of necrosis (Figure 2B,C).Immunohistochemically, the tumor cells showed strong positivity for pankeratin, EMA, and SOX10 (Figure 3).The tumor cells also exhibited patchy positivity for smooth muscle actin, F I G U R E 1 Representative magnetic resonance images of the tumor prior to neoadjuvant chemoradiation.Images show nodular tumor within the dermis and subcutaneous tissue on the plantar aspect to the first metatarsophalangeal joint of the right foot.(A) Sagittal short tau inversion recovery (STIR) image.(B) Axial STIR image.F I G U R E 2 Representative histological images of primary cutaneous nuclear protein in testis (NUT) carcinoma with BRD4::NUTM1 fusion.(A) Tumor cells are arranged in nodular aggregates within the dermis (H&E, Â20).(B) Tumor nest with apoptotic cells and necrosis (H&E, Â100).(C) Tumor cells have large vesicular and pleomorphic nuclei, prominent nucleoli, visible mitotic figures, and moderate amphophilic cytoplasm (H&E, Â200).(D) Lymphovascular invasion (H&E, Â200).

F
I G U R E 3 Representative immunohistochemical images of primary cutaneous nuclear protein in testis (NUT) carcinoma with BRD4::NUTM1 fusion.(A) Strong nuclear positivity for SOX10 immunohistochemistry in majority of tumor cells (Â200).(B) Cytoplasmic positivity for pancytokeratin immunohistochemistry (Â100).(C) Strong nuclear positivity for NUT immunohistochemistry in the majority of tumor cells (Â40).T A B L E 1 Clinicopathologic correlation of primary cutaneous nuclear protein in testis (NUT) carcinomas with corresponding fusions.

NSD3
rearrangements by showing a diffuse, poorly differentiated, and high-grade morphology as classically associated with NUT carcinomas from extracutaneous sites.Of note, previous studies on extracutaneous NUT carcinomas have shown that non-BRD4::NUT fusions confer a significantly better prognosis with a longer overall median F I G U R E 4 Schematic illustration of BRD4::NUTM1 fusion.Illustration of DNA translocation (top panel) resulting in the BRD4-NUT fusion protein (bottom panel).Courtesy of St. Jude PeCan.