Cutaneous rhabdomyosarcoma with FUS::TFCP2 fusion: A case report emphasizing early detection.

Rhabdomyosarcoma with TFCP2 rearrangement is a recently identified malignant neoplasm characterized by immunohistochemical evidence of rhabdomyoblastic differentiation, keratin expression, upregulation of ALK, and an aggressive clinical course. This neoplasm has a tendency to affect craniofacial bones, with only a few reported cases of extra-osseous tumors. Here, we present a case of cutaneous rhabdomyosarcoma with FUS::TFCP2 fusion in a 35-year-old female. Notably, the tumor exhibited a pathologic spectrum, initially resembling sclerosing dermatitis at presentation but progressing into a high-grade malignant tumor within 8 months. The distinctive immunoprofile of this neoplasm highlights the importance of early molecular studies for diagnosis, even in the presence of low-grade cytomorphology. Early detection may offer an opportunity for timely resection before the tumor becomes unresectable.

Epithelioid Rhabdomyosarcoma at the High Parietal Area of the Head: A Case Report.

Epithelioid rhabdomyosarcoma is a rare condition, which may be clinically misinterpreted as melanoma due to its morphological appearance. Careful morphological and immunohistochemical analysis play an important role in its diagnosis. This case report describes the clinicopathological features of an epithelioid RMS diagnosed at the high parietal area of the head. A 71-year-old male patient presented a red-brown pigmented ulcerative nodule in the high parietal region of the head. Previous biopsy and computer tomography imaging revealed a malignant melanoma in stage I (pT2, sN0, Mx). After tumor operation, histological and immunohistochemical analysis of the tumor were conducted. Histological analysis showed an erosive lesion with a monomorphic cell population containing small cells with prominent nucleoli. A positivity was confirmed for CD10, Vimentin, and Desmin. MyoD1 was detected, as well as a fluctuating signal for p53. Molecular analysis revealed a negativity for Sox-10, and a weak positivity for CK8/18 by absence of p40. Based on the morphological and immunohistochemical findings, the tumor was diagnosed as epithelioid RMS.

Recurrent VGLL3 fusions define a distinctive subset of spindle cell rhabdomyosarcoma with an indolent clinical course and striking predilection for the head and neck.

The mammalian Vestigial-like (VGLL) transcriptional cofactor family of proteins VGLL1-4 has recently emerged as an important player in the tumorigenesis of diverse neoplasms. The role of VGLL3 in soft tissue tumors is exemplified by its amplification in myxoinflammatory fibroblastic sarcoma and its rearrangement (fused to CHD7, CHD9, or MAMLD1) in hybrid schwannoma-perineurioma. This study characterizes a distinctive low-grade myogenic neoplasm with a striking predilection for the head and neck, characterized by VGLL3 fusions. The study includes five males and one female patient, aged 30-71 years (median, 56). Three tumors originated in the tongue, with one case each in the nasopharynx, oral cavity, and oropharynx. The VGLL3 fusion partners included TCF12 (n = 3), EP300 (n = 2), and PPARGC1A (n = 1). The tumor size range was 0.8-1.6 cm (all, but one, was <1 cm). Histologically, all tumors displayed bland spindle to ovoid cells arranged into vague fascicular and diffuse patterns. Mitotic activity ranged from 1 to 7 per 10 HPFs. Five tumors were muscle-centered and infiltrative, and one was centered beneath nasopharyngeal mucosa. Immunohistochemistry revealed consistent expression of desmin (diffuse in four and patchy in two cases) associated with patchy smooth muscle actin expression (4/6), and focal reactivity for myogenin (5/6) and myoD1 (1/3). All patients were managed surgically; one patient each received adjuvant radio- or chemotherapy. Three patients with follow-up were without disease at 8, 19, and 60 months and one was alive with unknown disease status at 24 months. All VGLL3 fusions were in-frame and involved exon 2, fused with either TCF12 exon 16, EP300 exon 31, or PPARGC1A exon 5, respectively. This series characterizes a distinctive subset of spindle cell rhabdomyosarcoma (RMS) with a predilection for the head and neck in adults, defined by VGLL3 fusions, likely indolent behavior and limited rhabdomyoblastic differentiation. Further delineation of this entity and differentiation from more aggressive molecular subtypes of spindle cell RMS is mandatory to define the most appropriate therapeutic strategy and avoid overtreatment.

ZFP64::NCOA3 gene fusion defines a novel subset of spindle cell rhabdomyosarcoma.

Spindle cell rhabdomyosarcoma represents a rare neoplasm characterized by monomorphic spindle cells with a fascicular architecture and variable skeletal muscle differentiation. Following incidental identification of a ZFP64::NCOA3 gene fusion in an unclassified spindle cell sarcoma resembling adult-type fibrosarcoma, we performed a retrospective archival review and identified four additional cases with a similar histology and identical gene fusion. All tumors arose in adult males (28-71 years). The neoplasms were found in the deep soft tissues, two were gluteal, and one each arose in the thigh, abdominal wall, and chest wall. Morphologically, the tumors were characterized by spindle cells with a distinctive herringbone pattern and variable collagenous to myxoid stroma. The nuclei were relatively monomorphic with variable mitotic activity. Three tumors had immunoreactivity for MyoD1, and four contained variable expression of desmin and smooth muscle actin. All cases tested for myogenin, CD34, S100, pankeratin, and epithelial membrane antigen were negative. Targeted RNA sequencing revealed a ZFP64::NCOA3 fusion product in all five tumors. Three patients developed distant metastases, and two ultimately succumbed to their disease within 2 years of initial diagnosis. This study suggests ZFP64::NCOA3 fusions define a novel subtype of rhabdomyosarcoma with a spindle cell morphology and aggressive clinical behavior. The potential for morphologic and immunohistochemical overlap with several other sarcoma types underscores the value of molecular testing as a diagnostic adjunct to ensure accurate classification and management of these neoplasms.

A subset of epithelioid and spindle cell rhabdomyosarcomas is associated with TFCP2 fusions and common ALK upregulation.

Rhabdomyosarcomas with TFCP2 fusions represent an emerging subtype of tumors, initially discovered by RNA-sequencing. We report herein the clinicopathological, transcriptional, and genomic features of a series of 14 cases. Cases were retrospectively and prospectively recruited and studied by immunohistochemistry (MYF4, MYOD1, S100, AE1/E3, ALK), fluorescence in situ hybridization with TFCP2 break-apart probe (n = 10/14), array-comparative genomic hybridization (Agilent), whole RNA-sequencing (Truseq Exome, Illumina), or anchored multiplex PCR-based targeted next-generation sequencing (Archer® FusionPlex® Sarcoma kit). Patient's age ranged between 11 and 86 years, including 5 pediatric cases. Tumors were located in the bone (n = 12/14) and soft tissue (n = 2/14). Most bone tumors invaded surrounding soft tissue. Craniofacial bones were over-represented (n = 8/12). Median survival was 8 months and five patients are currently alive with a median follow-up of 20 months. Most tumors displayed a mixed spindle cell and epithelioid pattern with frequent vesicular nuclei. All tumors expressed keratins and showed a rhabdomyogenic phenotype (defined as expression of MYF4 and/or MYOD1). ALK was overexpressed in all but three cases without underlying ALK fusion on break-apart FISH (n = 5) nor next-generation sequencing (n = 14). ALK upregulation was frequently associated with an internal deletion at genomic level. TFCP2 was fused in 5' either to EWSR1 (n = 6) or FUS (n = 8). EWSR1 was involved in both soft tissue cases. FISH with TFCP2 break-apart probe was positive in all tested cases (n = 8), including one case with unbalanced signal. On array-CGH, all tested tumors displayed complex genetic profiles with genomic indexes ranging from 13 to 107.55 and recurrent CDKN2A deletions. FET-TFCP2 rhabdomyosarcomas clustered together and distinctly from other rhabdomyosarcomas subgroups. Altogether, our data confirm and expand the spectrum of the new family of FET-TFCP2 rhabdomyosarcomas, which are associated with a predilection for the craniofacial bones, an aggressive course, and recurrent pathological features. Their association with ALK overexpression might represent a therapeutic vulnerability.

Beyond "Triton": Malignant Peripheral Nerve Sheath Tumors With Complete Heterologous Rhabdomyoblastic Differentiation Mimicking Spindle Cell Rhabdomyosarcoma.

Spindle cell rhabdomyosarcoma (RMS) is an aggressive sarcoma type with a predilection for the head and neck and frequent transactivating MYOD1 mutations. Malignant peripheral nerve sheath tumors (MPNST) show heterologous (most often rhabdomyoblastic) differentiation in 10% of cases; such tumors have been referred to as malignant "Triton" tumors. MPNST frequently harbors inactivating mutations in SUZ12 or EED, resulting in PRC2 dysfunction and loss of histone H3 lysine 27 trimethylation (H3K27me3), most often seen in sporadic and radiation-associated, high-grade tumors; immunohistochemistry (IHC) for H3K27me3 is a useful diagnostic marker. We recently encountered a tumor showing H3K27me3 loss but with otherwise typical features of spindle cell RMS. The purpose of this study was to evaluate H3K27me3 in spindle cell RMS and further investigate putative spindle cell RMS with loss of H3K27me3. IHC for H3K27me3 was performed on 50 tumors diagnosed as spindle cell RMS. Targeted sequencing of all exonic and selected intronic regions of ~450 genes was performed on the tumors with H3K27me3 loss using hybrid capture with a custom probe set and massively parallel (next-generation) sequencing (NGS). Of the 50 patients, 32 were male and 18 were female with a median age of 33 years (range, 6 wk to 77 y). Tumors most often involved head and neck (N=23), extremities/limb girdles (N=11), and trunk wall (N=5). Three cases (6%) showed loss of H3K27me3; based on all available evidence, we believe at least 2 of these cases in fact represent MPNST with complete heterologous rhabdomyoblastic differentiation: a deep-seated groin mass in a 76-year-old female and a paratesticular mass in a 22-year-old male (neither of whom had a history or signs of type 1 neurofibromatosis). The tumors showed similar histologic appearances: fascicular architecture, marked nuclear atypia, eosinophilic cytoplasm, and a high mitotic rate; rhabdomyoblasts were not apparent. One tumor showed focal areas with scant myxoid stroma and alternating hypocellularity and hypercellularity. By IHC, the tumors showed diffuse staining for desmin, myogenin, and MyoD1, whereas S100 protein and SOX10 were negative. NGS on 2 tumors revealed (1) 2-copy deletion of NF1, CDKN2A, and SUZ12 and a TP53 mutation with arm-level loss of 17p; and (2) 2-copy deletion of CDKN2A and an NF1 mutation with loss of 17q11, findings characteristic of MPNST. NGS on the third tumor showed no distinctive alterations. MPNST may occasionally show complete heterologous rhabdomyoblastic differentiation without histologic evidence of residual conventional MPNST, closely mimicking spindle cell RMS. IHC for H3K27me3 reliably distinguishes MPNST from spindle cell RMS.

Undifferentiated Sarcoma as Intermediate Step in the Progression of Malignant Melanoma to Rhabdomyosarcoma: Histologic, Immunohistochemical, and Molecular Studies of a New Case of Malignant Melanoma With Rhabdomyosarcomatous Differentiation.

Malignant melanoma (MM) may display highly variable phenotypic diversity, sometimes associated with loss of immunohistochemical melanocytic markers and acquisition of nonmelanocytic lineage of differentiation. Primary cutaneous MM with rhabdomyosarcomatous differentiation is extremely rare with only 5 reported cases in the literature. To date, a chronological progression of a MM to rhabdomyosarcoma has not been conclusively documented. A 96-year-old man underwent a re-excision of an "atypical fibroxanthoma" of the forearm, which revealed a small lentigo maligna melanoma associated with a dominant dermal high-grade spindle cell nodule admixed with a population of malignant polygonal epithelioid cells. On immunohistochemical studies, the spindle cells were completely negative for all melanocytic markers, whereas a small population of polygonal neoplastic cells at the periphery was positive for Desmin and Myo-D1, supporting early rhabdomyosarcomatous transformation. Several subsequent re-excisions demonstrated merely nodules of malignant pleomorphic epithelioid cells with rhabdomyosarcomatous differentiation and devoid of melanocytic markers. In addition, both rhabdomyosarcomatous component and original MM displayed identical mutations. Therefore, the histologic, immunohistochemical, and molecular findings documented for the first time a chronological progression from an invasive MM to a pleomorphic rhabdomyosarcoma through an intermediate stage of undifferentiated sarcoma/atypical fibroxanthoma. Interestingly, subsequent recurrences of pure rhabdomyosarcomatous component displayed skip lesions/microsatellitosis, marked tumor-infiltrative lymphocytes, and rare junctional nests of rhabdomyosarcomatous cells in the epidermis, histologic features that were not described in primary cutaneous rhabdomyosarcoma and therefore could serve as morphologic clues to the diagnosis of rhabdomyosarcomatous transformation in an MM.

Assessment of programmed death-ligand 1 expression and tumor-associated immune cells in pediatric cancer tissues.

BACKGROUND: Programmed death 1 (PD-1) signaling in the tumor microenvironment dampens immune responses to cancer, and blocking this axis induces antitumor effects in several malignancies. Clinical studies of PD-1 blockade are only now being initiated in pediatric patients, and little is known regarding programmed death-ligand 1 (PD-L1) expression in common childhood cancers. The authors characterized PD-L1 expression and tumor-associated immune cells (TAICs) (lymphocytes and macrophages) in common pediatric cancers. METHODS: Whole slide sections and tissue microarrays were evaluated by immunohistochemistry for PD-L1 expression and for the presence of TAICs. TAICs were also screened for PD-L1 expression. RESULTS: Thirty-nine of 451 evaluable tumors (9%) expressed PD-L1 in at least 1% of tumor cells. The highest frequency histotypes comprised Burkitt lymphoma (80%; 8 of 10 tumors), glioblastoma multiforme (36%; 5 of 14 tumors), and neuroblastoma (14%; 17 of 118 tumors). PD-L1 staining was associated with inferior survival among patients with neuroblastoma (P = .004). Seventy-four percent of tumors contained lymphocytes and/or macrophages. Macrophages were significantly more likely to be identified in PD-L1-positive versus PD-L1-negative tumors (P < .001). CONCLUSIONS: A subset of diagnostic pediatric cancers exhibit PD-L1 expression, whereas a much larger fraction demonstrates infiltration with tumor-associated lymphocytes. PD-L1 expression may be a biomarker for poor outcome in neuroblastoma. Further preclinical and clinical investigation will define the predictive nature of PD-L1 expression in childhood cancers both at diagnosis and after exposure to chemoradiotherapy. Cancer 2017;123:3807-3815. © 2017 American Cancer Society.

Diagnostic utility of cyclin D1 in the diagnosis of small round blue cell tumors in children and adolescents.

Small round blue cell tumors (SRBCTs) of children and adolescents are often diagnostically challenging lesions. With the increasing diagnostic approach based on small biopsies, there is the need of specific immunomarkers that can help in the differential diagnosis among the different tumor histotypes to assure the patient a correct diagnosis for proper treatment. Based on our recent studies showing cyclin D1 overexpression in both Ewing sarcoma/primitive peripheral neuroectodermal tumor (EWS/pPNET) and peripheral neuroblastic tumors (neuroblastoma and ganglioneuroblastoma), we immunohistochemically assessed cyclin D1 immunoreactivity in 128 cases of SRBCTs in children and adolescents to establish its potential utility in the differential diagnosis. All cases of EWS/pPNET and the undifferentiated/poorly differentiated neuroblastomatous component of all peripheral neuroblastic tumors exhibited strong and diffuse nuclear staining (>50% of neoplastic cells) for cyclin D1. In contrast, this marker was absent from rhabdomyosarcoma (regardless of subtype) and lymphoblastic lymphoma (either B- or T-cell precursors), whereas it was only focally detected (<5% of neoplastic cells) in some cases of Wilms tumor (blastemal component) and desmoplastic small round cell tumor. Our findings suggest that cyclin D1 can be exploitable as a diagnostic adjunct to conventional markers in confirming the diagnosis of EWS/pPNET or neuroblastoma/ganglioneuroblastoma. Its use in routine practice may also be helpful for those cases of SRBCT with undifferentiated morphology that are difficult to diagnose after application of the conventional markers.

Clinicopathologic analysis of spindle cell/sclerosing rhabdomyosarcoma.

BACKGROUND: Clinical characteristics and optimal treatment strategies for spindle cell/sclerosing rhabdomyosarcoma (ssRMS) have not been well established because of its rarity. PROCEDURE: Retrospective re-evaluation of sarcoma specimens (1997-2014) identified 16 ssRMSs (median age 20 years, range 7-39 years). Clinicopathological features, clinical course, and outcome were analyzed. RESULTS: Primary disease sites were the head and neck (10 cases) and other regions (6 cases). Nine cases were at Intergroup Rhabdomyosarcoma Study preoperative stage 3. The primary tumors were >5 cm in 13 cases. Two patients had lymph node metastases, but none had distant metastases at presentation. At follow-up (median period 39 months, range 4.6-201), seven patients were alive without disease. Among nine patients treated with the vincristine, actinomycin, and cyclophosphamide (VAC) regimen, five responded well, with four surviving free of disease. Among ten patients with recurrent or progressive disease, three experienced local recurrence, four had distant metastases, and three had both. None exhibited bone marrow invasion. Eight of the ten patients died in median time from relapse to death of 18 months (range 11-56). CONCLUSIONS: Although most ssRMSs present as a bulky tumor, nodal or distant metastases are rare at presentation. ssRMSs initially show good response to VAC, but >50% of tumors recur or progress; these data suggest a worse prognosis of ssRMS compared to the pediatric embryonal variant. As relapse typically occurs as local or distant solitary lesion without bone marrow invasion, localized treatment combined with chemotherapy would contribute to improve the prognosis of recurrent ssRMS.