GLIS Rearrangement is a Genomic Hallmark of Hyalinizing Trabecular Tumor of the Thyroid Gland

Background: Hyalinizing trabecular tumor (HTT) is a rare thyroid neoplasm with a characteristic trabecular growth pattern and hyalinization. This lesion has been the subject of long-term controversy surrounding its genetic mechanisms, relationship to papillary thyroid carcinoma (PTC), and malignant potential. Due to the presence of nuclear features shared with PTC, HTT frequently contributes to a false-positive cytology, which hampers patient management. The goal of this study was to apply genome-wide sequencing analyses to elucidate the genetic mechanisms of HTT and its relationship to PTC. Methods: Whole-exome, RNA-Seq, and targeted next-generation sequencing analyses were performed to discover and characterize driver mutations in HTT. RNA-Seq results were used for pathway analysis. Tissue expression of GLIS3 and other proteins was detected by immunohistochemistry. The prevalence of GLIS fusions was studied in 17 tumors initially diagnosed as HTT, 220 PTC, and 10,165 thyroid fine-needle aspiration samples. Results: Using whole-exome and RNA-Seq analyses of the initial three HTT, no known thyroid tumor mutations were identified, while in-frame gene fusion between PAX8 exon 2 and GLIS3 exon 3 was detected in all tumors. Further analysis identified PAX8-GLIS3 in 13/14 (93%) and PAX8-GLIS1 in 1/14 (7%) of HTT confirmed after blind pathology review. The fusions were validated by Sanger sequencing and FISH. The fusions resulted in overexpression of the 3′-portion of GLIS3 and GLIS1 mRNA containing intact DNA-binding domains of these transcription factors and upregulation of extracellular matrix genes including collagen IV. Immunohistochemistry confirmed upregulation and deposition of collagen IV and pan-collagen in HTT. The analysis of 220 PTC revealed no PAX8-GLIS3 and one PAX8-GLIS1 fusion. PAX8-GLIS3 was prospectively identified in 8/10,165 (0.1%) indeterminate cytology fine-needle aspiration samples; 5/5 resected fusion-positive nodules were HTT on surgical pathology. Conclusions: This study demonstrates that GLIS rearrangements, particularly PAX8-GLIS3, are highly prevalent in HTT but not in PTC. The fusions lead to overexpression of GLIS, upregulation of extracellular matrix genes, and deposition of collagens, which is a characteristic histopathologic feature of HTT. Due to unique genetic mechanisms and an indolent behavior, it is proposed to rename this tumor as "GLIS-rearranged hyalinizing trabecular adenoma."