Second Report of PDE10A-BRAF Fusion in Pediatric Spindle Cell Sarcoma With Infantile Fibrosarcoma-Like Morphology Suggesting PDE10A-BRAF Fusion Is a Recurrent Event

Abstract

Infantile/congenital fibrosarcoma (IFS) is the most common soft tissue tumor in children less than one year of age. The most common anatomic site of IFS is in the extremities or trunk, and rarely in the abdomen or retroperitoneum. Approximately 70–90% of cases are characterized by a distinct t(12;15)(p13;q25) translocation resulting in an ETV6-NTRK3 gene fusion. As such, TRK inhibitors are considered frontline therapy in TRK-fusion positive IFS. The ETV6-NTRK3 fusion is also detected in congenital mesoblastic nephroma (CMN) and less frequently in myeloid leukemias, secretory breast carcinoma, and mammary-type secretory carcinoma of the skin and salivary glands. Infrequently, cases of tumors with IFS-like morphology without the characteristic ETV6-NTRK3 gene fusion have been identified. Herein, an ETV6-NTRK3 fusion negative spindle cell sarcoma with IFS-like morphology subjected to genomic profiling revealed a PDE10A-BRAF fusion, a fusion event that has been detected previously in an isolated case of undifferentiated infantile sarcoma.

Introduction

Soft tissue sarcomas comprise between 8–10% of pediatric cancers of which 10% are classified as infantile/ congenital fibrosarcoma (IFS). IFS is the most common soft tissue tumor in children less than one year of age.¹ IFS occurs most commonly in the extremities or trunk, with rare tumors found within the abdomen or the retroperitoneum.² IFS exhibits a broad morphologic spectrum, from fascicles to sheets of spindled cells within a collagenous or myxoid stroma to a hemangiopericytoma-like vascular pattern with plump, rounded cells. This variation in morphology should prompt a wide differential diagnosis, inclusive of both benign and malignant entities such as infantile fibromatosis and infantile myofibroma/myofibromatosis, amongst others. Most cases of IFS (~70–90%) are characterized by a distinct t(12;15)(p13;q25) translocation resulting in an ETV6-NTRK3 gene fusion; rarely other partners are seen such as EML4-NTRK3.^1,3 The ETV6-NTRK3 gene fusion product functions as a constitutively active tyrosine kinase in the Ras-MAPK and PI3K-Akt pathways.⁴ The ETV6-NTRK3 fusion is not unique to IFS; it is also detected in congenital mesoblastic nephroma (CMN) and less frequently in myeloid leukemias, secretory breast carcinoma, and mammary-type secretory carcinoma of the skin and salivary glands.⁴ With sequencing being performed more frequently, the spectrum of tumors with ETV6-NTRK3 fusion is expanding and now includes radiation-associated thyroid cancer, a subset of sinonasal low-grade non-intestinal type adenocarcinoma, and some ALK-negative inflammatory myofibroblastic tumors.^5–7 Other fusions involving NTRK, MET, RAF1, BRAF genes or intragenic BRAF events have been reported in infantile fibrosarcoma or infantile fibrosarcoma-like tumors.^4,8–12 Such BRAF fusions or intragenic alterations result in the loss of the negative regulatory RAS-binding domain, leading to constitutive activation of BRAF and the MAP kinase (MAPK) signaling pathway.

IFS has an excellent prognosis, with overall survival rates of 80–94%.^13,14 Metastatic disease is uncommon. Cure can be achieved through complete surgical resection, though local recurrence is common with incomplete or margin-positive resection, which requires radiation or re-resection. Neoadjuvant chemotherapy is necessary in patients where complete surgical resection is not feasible. Historically, this treatment has consisted of multiagent chemotherapy with vincristine and actinomycin (VA) with or without cyclophosphamide.¹⁴ With the recognition that most IFS harbor NTRK gene fusions, and the growing use of TRK inhibitors in IFS and other TRK-fusion positive tumors, TRK inhibitors are rapidly becoming the frontline therapy in TRK-fusion positive IFS.^15–17

Case Report

A 5-month-old female presented to the emergency department with bilateral lower extremity paralysis, muscle wasting and constipation. The patient’s mother reported that her daughter had a small lump on the left side of her spine since birth (Figure 1(A)). Magnetic resonance imaging (MRI) demonstrated a 6.8 cm epidural mass compressing the spinal cord, displacing the thecal sac, with extension into the psoas muscle, and through the L1 and L2 foramina (Figure 1(B)). The patient was taken urgently to the operating room for a multilevel thoracolumbar laminectomy and debulking surgery.

Figure 1.

A, Left-sided posterior soft tissue mass measuring approximately 8 cm. B, Magnetic resonance imaging (MRI) showing 6.8 cm epidural mass (arrow) extending from the left L1 and L2 foramina, displacing the thecal sac.

Histological examination of the mass demonstrated monomorphic spindled cells arranged in a fascicular architecture with abundant mitotic figures (up to 27 per high-power field) (Figure 2). Immunohistochemistry demonstrated negative staining for NKX2.2, CD99, MyoD1, and myogenin. INI-1 expression was retained. Based on the clinical history and the morphologic features of the mass, FISH analysis for ETV6 rearrangement was performed and was negative. Additional testing was recommended to determine if this case harbored a variant translocation, alternative TRK fusions (NTRK1 or NTRK2), or a fusion/intragenic alteration involving other kinases that has been reported in a subset of IFS or IFS-like cases (10–30%).^4,8–12

Figure 2.

A, The hypercellular mass is composed of sheets of cells arranged in a vaguely fasciular architecture surrounded focally by a pseudocapsule (4×). B, The cells are spindled to ovoid with abundant mitotic figures, up to 27 per high-power field, and numerous hemangiopericytoma-like vessels scattered throughout (20×). C, On higher power, the spindled to ovoid cells are relatively uniform with inconspicuous nucleoli (40×).

Whole exome/whole transcriptome next-generation sequencing (NGS) was performed as previously described¹⁸ and demonstrated an in-frame fusion between exon 3 of the phosphodiesterase 10A (PDE10A) gene and exon 11 of the BRAF (B-Raf proto-oncogene, serine/threonine kinase) gene (Figure 3). This fusion gene event is predicted to result in the generation of a 403 amino acid protein that contains the tyrosine kinase catalytic domain of BRAF under the control of the PDE10A promoter. Subsequent FISH analysis revealed an abnormal pattern featuring one normally juxtaposed orange/green BRAF flanking probe signal accompanied by a single green signal (just 3′ to BRAF) consistent with a rearrangement of the BRAF (7q34) locus in 95% of the 100 interphase nuclei analyzed (Figure 4). Specifically, an isolated 3′BRAF probe signal supports the findings of the PDE10A-BRAF fusion identified by NGS whereby only exons 11–17 containing the kinase domain at the 3′ C-terminal end of BRAF are part of the fusion event.

Figure 3.

Schematic gene structure of PDE10A and BRAF with resulting novel gene fusion product.

Figure 4.

Custom BRAF breakapart probe FISH study with green-labeled probe located just 3′ to BRAF and orange-labeled probe located 5′ to BRAF demonstrating an aberrant pattern of an isolated 3′ BRAF (green) signal accompanying a normally juxtaposed orange/green signal in the majority of interphase nuclei evaluated consistent with an unbalanced rearrangement of the BRAF locus.

The patient’s post-operative course was complicated by wound dehiscence and infection, delaying initiation of chemotherapy. MRI imaging showed substantial interval regrowth of the tumor in just five weeks after the initial debulking procedure. The patient was then started on a chemotherapy regimen of vincristine, actinomycin-d and cyclophosphamide (VAC). She completed 14 cycles of chemotherapy with proton radiation to the primary tumor site during the final 6 weeks of treatment. After therapy completion, she has regained some function in her right lower extremity. She is now 5 months from completion of therapy and her end of therapy imaging does not demonstrate any evidence of residual disease.

Discussion

This case illustrates the increasing importance of molecular testing in the diagnosis of pediatric tumors.¹⁹ For rare tumors like IFS where an ETV6-NTRK3 fusion is suspected, NTRK3 FISH should be performed to assess the utility of highly effective TRK inhibitors, such as entrectinib and larotrectinib.²⁰ If FISH is negative, other profiling studies such as NGS should be explored as the molecular heterogeneity of tumors that were previously thought to be defined by a single driver event, such as ETV6-NTRK3 gene fusion in IFS, is being uncovered.

This unique PDE10A-BRAF fusion has been described previously in an isolated case of a 7.5 cm undifferentiated sarcoma of the pelvis in a 3-month-old female composed of spindled cells with clear cytoplasm and a high mitotic rate.²¹ Initial workup of this 3-month-old revealed a complex karyotype but no rearrangements of SS18, ETV6, DUX4, nor CCNB3. The patient received chemotherapy with five cycles of doxorubicin and six cycles of ifosfamide as surgical resection was deemed to be too high risk for neurologic sequelae given the location of the tumor. After 18 months, there was progression of the residual mass. RNA sequencing analysis was performed, revealing an in-frame fusion between exon 3 of PDE10A and exon 9 of BRAF. After local recurrence following frontline chemotherapy, the 3-month-old then responded to trametinib, a MEK (MAPK pathway) inhibitor.²¹ To compare, in the case described herein, the 5-month-old was treated with surgical resection followed by fourteen cycles of the historical standard of vincristine, actinomycin-d and cyclophosphamide (VAC) in addition to proton radiation to the primary tumor site during the final 6 weeks of treatment.¹⁴ Targeted MAPK therapy would have been considered in this case if the patient had relapsed.

Other BRAF fusions have been identified in unclassified spindle cell sarcomas with morphologic resemblance to IFS.^8,10,19,21 Kao et al. reports five IFS-like pediatric sarcomas without the hallmark ETV6-NTRK3 fusion that were found to have alternative BRAF gene fusions. Four of the five were located in intra-abdominal and retroperitoneal sites; the fifth was located in the thigh. Reported fusion partners include CUX1 and SEPT7.⁸ In addition, Wegert et al. identified three cases of IFS with intragenic BRAF deletions, with two of the three co-occurring with the characteristic ETV6-NTRK3 fusion.¹⁰ BRAF fusions or intragenic alterations result in the loss of the negative regulatory RAS-binding domain, leading to constitutive activation of BRAF and the MAPK signaling pathway.

Including this case, a total of seven cases of pediatric spindle cell sarcomas with BRAF gene fusions have been reported.^8,21 Additionally, three intragenic BRAF deletions have been identified in tumors with IFS-like morphology, of which the anatomic sites were not reported. Of the masses with known and documented anatomic sites, six of the seven (6/7) tumors were located within the abdomen or retroperitoneum. These cases suggest the possibility that a distinct subset of IFS-like tumors with BRAF gene fusions have a predilection for the abdomen/retroperitoneum and may represent a separate pediatric sarcoma entity entirely. As such, pediatric spindle cell sarcomas with IFS-like morphology that do not have the hallmark ETV6-NTRK3 should be queried for alternative gene fusions by NGS, especially when they are in the abdomen/retroperitoneum. Further research is needed to determine whether BRAF-fusion positive IFS have different outcomes than TRK-fusion positive IFS, and whether targeting the MAPK pathway with tyrosine kinase inhibitors (TKIs) might achieve the same robust responses that TRK inhibitors do for TRK-fusion positive IFS. If so, the discovery of these recurrent BRAF fusions may help spare these infants from the toxicity and late side effects of conventional chemotherapy.