ASXL1 and DNMT3A mutation in a cytogenetically normal B3 thymoma

Abstract

The molecular drivers of thymoma are poorly understood. Outside of the identification of rarely occurring epidermal growth factor receptor and v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog mutations via candidate gene sequencing, mutations in common cancer genes have yet to be observed. Only a single thymoma genome sequence has been previously reported, with no mutations in known cancer genes identified. Thus, we attempted to identify somatic driver mutations in a cytogenetically normal thymoma. A stage IVB type B3 thymoma from a 47-year-old male of Asian descent with no history of myasthenia gravis or other autoimmune condition was genomically evaluated. Exome sequencing and low-pass whole-genome sequencing was performed to identify somatic point mutations, copy number changes and structural variants. Mutations in known tumor suppressors DNMT3A (p.G728D) and ASXL1 (p.E657fs), consistent with mutations of known consequence in acute myeloid leukemia, were identified. Contrary to a previous report, this finding suggests the genetic etiology of thymomas may not be fundamentally distinct from other tumor types. Rather, these findings suggest that further sequencing of cytogenetically normal thymoma samples should reveal the specific molecular drivers of thymoma.

Figure 1

(a) Thymoma imaging. fluorodeoxyglucose positron emission tomography/computed tomography (CT) demonstrates intensely hypermetabolic anterior mediastinal mass on fused coronal and axial images (arrowheads). Cystic tumor component on attenuation-correction CT image is non-hypermetabolic on FDG PET (arrows). (b) Renal tumor imaging. CT scan with contrast (left) demonstrates large contrast-enhancing left renal mass (arrowheads) with posterior hydronephrotic upper pole collecting system (block arrow). Fused axial FDG PET/CT (right) demonstrates mildly hypermetabolic posterior tumor component (arrow). Upper pole hydronephrosis contains excreted urinary FDG (block arrow). (c, d) Thymoma histology. (c) Lobules of thymoma tumor cells infiltrate into the adjacent lung parenchyma (arrows: hematoxylin and eosin (H&E): × 5 magnification). (d) Sheets of thymoma type B3 polygonal tumor cells with ovoid nuclei (H&E: × 20 magnification). (e) Tumor genome overview. Circos plot overview of somatic mutations identified in the thymoma. Outer text denotes location and gene impacted by somatic protein coding mutations. Middle gray ring depicts copy number variation (CNV) status (black=2 copies, red=CNV gains, blue=CNV losses). Inner text displays genes overlapping identified CNVs. Inner red lines display identified structural variants.

Figure 2

The DNMT3A p.G728D mutation observed in this thymoma sample is visualized in the context of other DNMT3A mutations observed in AML genome sequences from TCGA. The protein sequence and functional domains are depicted on the x axis. The number of AML mutations is depicted on the y axis. Red circles correspond to truncating mutations. Green circles correspond to missense mutations. Circle height corresponds to the number of mutations per position, however, the G728D indicator (black) is only meant to indicate position of this mutation. Note the clustering of AML nonsynonymous mutations around position 728. Mutations in this region reduce DNMT3A activity by disrupting the interaction between DNMT3L and DNMT3A. The Sanger sequencing validation trace of p.G7238D is also shown, demonstrating validation of p.G7238D as a homozygous somatic mutation.

Figure 3

The ASXL1 p.E657fs mutation observed in this thymoma sample is visualized in the context of other ASXL1 mutations observed in all tumor genome sequences catalogued in TCGA. The protein sequence and functional domains are depicted on the x axis. The number of TCGA mutations is depicted on the y axis. Red circles correspond to truncating mutations. Green circles correspond to nonsynonymous mutations. Purple circles correspond to mutations that are both nonsynonymous and truncating in different gene isoforms. Circle height corresponds to the number of mutations per position, however, the E657fs indicator (black) is only meant to indicate position of this mutation. Note the clustering of TCGA truncating mutations around position 657. The Sanger sequencing validation trace of p.E657fs is also shown, demonstrating validation of p.E657fs as a heterozygous somatic mutation.