Molecular Profiling of Sinonasal Adenoid Cystic Carcinoma: Canonical and Noncanonical Gene Fusions and Mutation

Abstract

Adenoid cystic carcinomas (AdCC) of salivary gland origin have long been categorized as fusion-defined carcinomas owing to the almost universal presence of the gene fusion MYB::NFIB , or less commonly MYBL1::NFIB. Sinonasal AdCC is an aggressive salivary gland malignancy with no effective systemic therapy. Therefore, it is urgent to search for potentially targetable genetic alterations associated with AdCC. We have searched the authors' registries and selected all AdCCs arising in the sinonasal tract. The tumors were examined histologically, immunohistochemically, by next generation sequencing (NGS) and/or fluorescence in situ hybridization (FISH) looking for MYB/MYBL1 and/or NFIB gene fusions or any novel gene fusions and/or mutations. In addition, all tumors were tested for HPV by genotyping using (q)PCR. Our cohort comprised 88 cases of sinonasal AdCC, predominantly characterized by canonical MYB::NFIB (49 cases) and MYBL1::NFIB (9 cases) fusions. In addition, noncanonical fusions EWSR1::MYB ; ACTB::MYB; ESRRG::DNM3 , and ACTN4::MYB were identified by NGS, each of them in 1 case. Among nine fusion-negative AdCCs, FISH detected rearrangements in MYB (7 cases) , NFIB (1 case), and EWSR1 (1 case). Six AdCCs lacked fusions or gene rearrangements, while 11 cases were unanalyzable. Mutational analysis was performed by NGS in 31/88 (35%) AdCCs. Mutations in genes with established roles in oncogenesis were identified in 21/31 tumors (68%), including BCOR (4/21; 19%), NOTCH1 (3/21; 14%), EP300 (3/21; 14%), SMARCA4 (2/21; 9%), RUNX1 (2/21; 9%), KDM6A (2/21; 9%), SPEN (2/21; 9%), and RIT1, MGA, RB1, PHF6, PTEN, CREBBP, DDX41, CHD2, ROS1, TAF1, CCD1, NF1, PALB2, AVCR1B, ARID1A, PPM1D, LZTR1, GEN1 , PDGFRA , each in 1 case (1/21; 5%). Additional 24 cases exhibited a spectrum of gene mutations of uncertain pathogenetic significance. No morphologic differences were observed between AdCCs with MYBL1::NFIB and MYB::NFIB fusions. Interestingly, mutations in the NOTCH genes were seen in connection with both canonical and noncanonical fusions, and often associated with high-grade histology or metatypical phenotype, as well as with poorer clinical outcome. Noncanonical fusions were predominantly observed in metatypical AdCCs. These findings emphasize the value of comprehensive molecular profiling in correlating morphologic characteristics, genetic landscape, and clinical behavior in AdCC.

FIGURE 1

AdCC with canonical MYB:NFIB gene fusion showed biphasic tumor growth of predominantly cribriform pattern (A) than tubular or more solid areas (B). Both pseudocysts and true glandular lumina were characteristic for AdCC (C). Abundant deposits of a hyalinized eosinophilic extracellular matrix and reduplicated basement membrane materials were often seen (D). The fusion joining of MYB gene exon 14 with NFIB gene exon 9 is illustrated. Protein domains are depicted (E).

FIGURE 2

AdCC with canonical MYBL1:NFIB gene fusion was composed predominantly of combination of cribriform (A) and tubular (B) patters with microcystic-like spaces filled with hyaline or basophilic basement membrane material (C). The solid areas were also present lacking pseudocystic spaces present in previous 2 patters (D). The fusion joining of MYBL1 gene exon 14 with NFIB gene exon 9 is illustrated. Protein domains are depicted (E).

FIGURE 3

The case of ACTBex3:MYBex3 AdCC the tumor resemble myoepithelial derived neoplasm with hypercellular periphery and necrotic center (A). The tumor cells were basaloid with monomorphic hyperchromatic nuclei (B) and areas with high-grade features consisting of moderately pleomorphic nuclei (C). Conventional cribriform or tubular patterns were present at the periphery of the tumor surrounded by fibrohyaline stroma (D).

FIGURE 4

A, The AdCC with in-frame ACTN4ex18:MYBex2 gene fusion consisted of 2 types of tumor cells with predominant monomorphic and basaloid cells and groups of vacuolated cells. B, Stromal component was minor and characterized by mild cellular fibrous bands with distended vascular spaces dispersed throughout the tumor. C, Basaloid tumor cells were positive for p63. D, Vacuolated (clear) cells showed CK7 expression.

FIGURE 5

A, The case of ESRRGex3:DNM3ex14 translocated AdCC consisted of different architectural patterns including solid, tubular, micropapillary, and cribriform. B, Tumor cells were pleomorphic with visible nucleoli, nuclear hyperchromasia, and focally vacuolated cytoplasm. C, Aggressive appearance of the tumor was supported bymassive targetoid perineural spread. D, A transition zone contained atypical sinonasal glands arising from the respiratory adenomatoid hamartoma.

FIGURE 6

The AdCC with in-frame EWSR1ex6:MYBex2 gene fusion was in part composed of cellular areas of basaloid cells forming tubular and cribriform patterns with multiple areas of comedo-like necrosis (A), focally with extensive tubular hypereosinophilia (B) and foci of intraluminal clear cells (C) separated by fibrous bands. In part, the tumor consisted of hypocellular characterized by fibrosclerotic and hyalinized stroma with focal myxoid change and cords and ducts of tumor cells (D).

FIGURE 7

A, The cases of AdCC with NOTCH1/3 mutation were predominantly of solid growth focally with pseudocysts filled with basophilic mucoid material. B, The solid areas showed comedonecrosis and were embedded in desmoplastic stroma with bone invasion. C, Conventional cribriform or tubular patterns were less encountered. D, Ki-67 index was high in solid areas.

FIGURE 8

The metatypical changes were encountered in 2 AdCC with NOTCH mutations showing squamous cell differentiation (A), tubular hypereosinophilia (B), focal sebaceous metaplasia (C), and foamy vacuolated cells (D).