RNA sequencing identifies a novel USP9X‐USP6 promoter swap gene fusion in a primary aneurysmal bone cyst

Abstract

Primary aneurysmal bone cyst (ABC) is a benign multiloculated cystic lesion of bone that is defined cytogenetically by USP6 gene rearrangements. Rearrangements involving USP6 are promoter swaps, usually generated by fusion of the noncoding upstream exons of different partner genes with exon 1 or 2 of USP6, thus leading to transcriptional upregulation of full‐length USP6 coding sequence. Testing for USP6 rearrangements is used diagnostically to distinguish it from secondary ABC and other giant cell‐rich primary bone tumors. In this report, we present a case of a 16‐year‐old male with a primary ABC of the left distal femur. USP6 break apart fluorescence in situ hybridization was positive for a rearrangement and conventional chromosome analysis identified a reciprocal X;17 translocation. In order to identify the putative USP6 fusion partner, we performed RNA sequencing and uncovered a novel USP9X‐USP6 promoter swap fusion. This result was confirmed by reverse transcription‐polymerase chain reaction (RT‐PCR) and by mate pair sequencing thus showing the utility of these alternative methodologies in identifying novel fusion candidates. Ubiquitin‐specific protease 9X (USP9X), like USP6, encodes a highly conserved substrate‐specific deubiquitylating enzyme. USP9X is highly expressed in a number of tissue types and acts as both an oncogene and tumor suppressor in several human cancers. We conclude that oncogenic activation of USP6 via USP9X promoter exchange represents a novel driver of primary ABC formation.

Novel ASAP1‐USP6, FAT1‐USP6, SAR1A‐USP6, and TNC‐USP6 fusions in primary aneurysmal bone cyst

“…A specific fusion partner does not seem to influence the structure of the USP6 gene product, but rather the level of its overexpression. From our and other recent small series of ABCs, it is not possible to draw conclusions about the clinical or pathologic significance of different fusion partners . Large studies with longer follow‐up are needed to determine whether specific fusion partners lead to specific morphologic features of ABCs, are associated with particular locations in the skeleton or determine biologic behavior …”

“…NGS‐based analyses have not only shown the universal presence of USP6 gene rearrangement in primary ABCs, but also revealed several novel fusion partners. Hitherto, 14 different USP6 fusion partners have been identified: CDH11 , CNBP , COL1A1 , CTNNB1 , EIF1 , FOSL2 , OMD , PAFAH1B1 , RUNX2 , SEC31A , SPARC , STAT3 , THRAP3 and USP9X , with the CDH11‐USP6 fusion being the most common (reported in 19% and 38% of cases in two series) . In this study, we describe four novel fusion partners of the USP6 gene: FAT1 , TNC , ASAP1 , and SAR1A .…”

“…From our and other recent small series of ABCs, it is not possible to draw conclusions about the clinical or pathologic significance of different fusion partners. [3][4][5][8][9][10] Large studies with longer follow-up are needed to determine whether specific fusion partners lead to specific morphologic features of ABCs, are associated with particular locations in the skeleton or determine biologic behavior. 2,3,5 USP6 gene rearrangements have been identified in tumors other than bone ABCs, including nodular fasciitis, 28,29 soft tissue ABCs, 9,30 fibro-osseous pseudotumor of digits 31,32 and myositis ossificans.…”

“…There is a growing list of known fusion partners, of which CDH11 seems to be the most frequent; it was detected in 19% and 38% of ABCs in two series . Other fusion partners identified so far include CNBP , COL1A1 , OMD and THRAP3 , CTNNB1 , E1F1 , FOSL2 , PAFAH1B1 , RUNX2 , SEC31A , STAT3 , SPARC and, most recently, USP9X . Despite different fusion partners, all USP6 rearrangements seem to function by a promoter swap mechanism, resulting in oncogenic activation of USP6 and subsequent ABC formation .…”

“…2,3 Other fusion partners identified so far include CNBP, COL1A1, OMD and THRAP3, CTNNB1, E1F1, FOSL2, PAFAH1B1, RUNX2, SEC31A, STAT3, SPARC and, most recently, USP9X. [3][4][5][8][9][10] Despite different fusion partners, all USP6 rearrangements seem to function by a promoter swap mechanism, resulting in oncogenic activation of USP6 and subsequent ABC formation. 3 As suggested by a recent study in which a USP6 fusion was detected in all 13 ABCs analyzed by NGS and 7 novel fusion partners were detected in 8 cases, it seem very likely that some USP6 fusion partners still remain undiscovered.…”