A comprehensive analysis of RHOA mutation positive and negative angioimmunoblastic T-cell�lymphomas by targeted deep sequencing, expression profiling and single cell digital image analysis

Abstract

Angioimmunoblastic T-cell lymphoma (AITL) is a uniquely aggressive mature T-cell neoplasm. In recent years, recurrent genetic mutations in ras homolog family member A ( RHOA ), tet methylcytosine dioxygenase 2 ( TET2 ), DNA methyltransferase 3 alpha ( DNMT3A ) and isocitrate dehydrogenase [NADP(+)] 2 ( IDH2 ) have been identified as associated with AITL. However, a deep molecular study assessing both DNA mutations and RNA expression profile combined with digital image analysis is lacking. The present study aimed to evaluate the significance of molecular and morphologic features by high resolution digital image analysis in several cases of AITL. To do so, a total of 18 separate tissues from 10 patients with AITL were collected and analyzed. The results identified recurrent mutations in RHOA , TET2 , DNMT3A , and IDH2 , and demonstrated increased DNA mutations in coding, promoter and CCCTC binding factor (CTCF) binding sites in RHOA mutated AITLs vs. RHOA non-mutated cases, as well as increased overall survival in RHOA mutated patients. In addition, single cell computational digital image analysis morphologically characterized RHOA mutated AITL cells as distinct from cells from RHOA mutation negative patients. Computational analysis of single cell morphological parameters revealed that RHOA mutated cells have decreased eccentricity (more circular) compared with RHOA non-mutated AITL cells. In conclusion, the results from the present study expand our understanding of AITL and demonstrate that there are specific cell biological and morphological manifestations of RHOA mutations in cases of AITL.

Metabolic determinants of tumour initiation

“…Supporting this model, IDH1/IDH2 and TET2 mutations are mutually exclusive in patients with acute myelogenous leukaemia 33 . However, IDH2 and TET2 mutations do co-occur in a subset of angioimmunoblastic T cell lymphomas, indicating that the dioxygenases responsible for the phenotypes downstream of d-2-hydroxyglutarate accumulation are likely to be context-specific 116,117 . Moreover, although TET2 mutations are frequently found in clonal haematopoiesis -a benign precursor of leukaemia -IDH1/IDH2 mutations are not, suggesting that the IDH1/IDH2 mutation is not a common initiating event in acute myelogenous leukaemia 118 .…”

Chidamide plus prednisone, etoposide, and thalidomide for untreated angioimmunoblastic T‐cell lymphoma in a Chinese population: A multicenter phase II trial

“…AITL has been associated with several gene mutations, 26 including recurrent genetic mutations in ras homolog family member A (RHOA, 50-79%), [27][28][29] tet methylcytosine dioxygenase 2 (TET2, 68-80%), [29][30][31][32] DNA methyltransferase 3 alpha (DNMT3A, 20-30%), 28,29 and isocitrate dehydrogenase [NADP(+)] 2 (IDH2, 20-30%), 28,29 which are regulated by acetylation. 33,34 Histone deacetylases (HDACs) act as epigenetic modifiers that regulate signaling pathways via the deacetylation of histone proteins and non-histone proteins.…”

Mutations in JAK/STAT and NOTCH1 Genes Are Enriched in Post-Transplant Lymphoproliferative Disorders

“…We extracted DNA from formalin fixed and paraffin embedded (FFPE) tissue using the DNA Storm Kit (Cell Data Science, CA, USA). For our DNA libraries, we developed a customized SureSelect XT HS (Agilent Technologies, CA, USA) Heme Malignancies Evaluation and Infectious Detection panel (HeME-ID), which includes 354 genes that are important for lympho- or leukemogenesis in addition to 13 viruses and bacteria associated with hematolymphoid diseases ( 9 , 10 ). 100 base-pair paired-end targeted deep sequencing was performed at an average coverage depth of 1500-fold on a high-throughput sequencing platform (HiSeq4000).…”