Novel Mutations in NOTCH and Altered Wnt/β-Catenin Pathway Indicate a Role of Embryonic Signals in the Pathogenesis of T-Cell Prolymphocytic Leukemia
Authors: Zhang Henan , Tian Shulan , Braggio Esteban , Feldman Andrew L , Ketterling Rhett P. , He Rong , Call Timothy G. , Witzig Thomas E. , Parikh Sameer A. , Lin Yi , Secreto Charla R , Andersson Emma I , Mustjoki Satu , Schrader Alexandra , Herling Marco , Yan Huihuang , Ding Wei 
Published: October 2019

Background:Recurrent mutations in ATM, JAK1/3 and STAT5B genes have been identified in T-cell prolymphocytic leukemia (T-PLL), a mature T cell leukemia with poor survival. However, much remains to be understood regarding the dysregulated pathways involved in the pathogenesis of this aggressive disease. Here, we integrated the analyses of whole exome sequencing (WES) and transcriptomes of a cohort of 12 T-PLL samples to gain further insights into molecular mechanisms of this malignancy.

Methods: Leukemic cells were isolated from peripheral blood or bone marrow samples of T-PLL patients. Tumor DNA and RNA, and germline DNA were extracted from purified leukemic T cells and bone marrow derived stromal cells, respectively. WES (n=12) and transcriptome analysis (n=10) were performed and data analyzed with published standard methods (Robinson et al. 2010 and DePristo et al. 2011)

Results: 7 paired tumor and germline DNA and 5 tumor-only DNA samples were used for WES analysis. Mutations in ATM and TET2 were detected in 4 (33%) and 1 (8%) patients, respectively. JAK3 (p.M511I, p.Q501H and p.Q503H) or STAT5B (2 p.T628S and 1 p.N642H) mutations each occurred in 3 (25%) cases and were mutually exclusive, consistent with prior reports. Importantly, we found several novel mutations in genes with regulatory roles in the NOTCH signaling pathway. Specifically, 2 somatic mutations p.A3V and p.A3S in NOTCH2 and 2 other mutations (p.S244L and p.A1414V) in NOTCH4 were identified in 4 (33%) patients. One mutation was in the EGF- like and another one in the NODP domain critical for NOTCH-mediated cellular differentiation. Two (16.7%) novel missense mutations (p.G174D and p.V836I) were found in MAML2. Three missense mutations (2 p.S583N and 1 p.R190H) occurred in MAML1. Both MAML2 and MAML1 are essential cofactors of the NOTCH signaling pathway. In addition, a somatic mutation in the NOTCH signal regulator DTX2 (p.G268D) was detected. Altogether 7 (58.3%) patients harbored mutations in genes with a regulatory role in the NOTCH pathway. Among the epigenetic modifiers, we found deletions and point mutations in KDM6B and KDM6A in 4 (30%) patients including 2 codon deletions in KDM6B (p.X751Val) and 1 missense mutation each in KDM6B (p.R1016Q) and KDM6A (p.V510G). KDM6A and KDM6B are histone H3 lysine 27 demethylases known to be involved in regulation of DNA repair and play an important role in embryonic differentiation.

Transcriptome analyses of 10 T-PLL cases versus normal CD3+ T cells from 5 healthy donors revealed 706 up-regulated and 655 down-regulated genes. Consistent with prior data, TCL1A and TCL1B were significantly overexpressed (23.7 and 13.1 fold, respectively). Using Ingenuity Pathway Analysis, we found PTEN Signaling, glutamate receptor, and axonal guidance signaling to be among the top 3 upregulated pathways. The top upstream regulators implicated for the up-regulated signals were FGF, Wnt/β-catenin and TNF. The top 3 down-regulated pathways were granulocyte adhesion and diapedesis, IL-17A in psoriasis and IL-10 signaling. The top regulators for the down-regulated pathways were LPS, TNF and IFNg. Wnt/β-catenin and Human embryonic stem cell pluripotency signaling were among the top 20 up-regulated pathways. Many genes involved in the regulation of Wnt signal pathway were included in the above pathways, including Wnt ligands (Wnt9A, Wnt4), transmembrane receptors (CDH1, LRP5, FZD6, FZD7, SMO), cytoplasmic factor (AXIN2), and nuclear factors (MYC, SOX12, SOX8, SOX18, CCND1).

Summary: Our study identified recurrent mutations in the NOTCH signaling pathway as well as dysregulated embryonic developmental pathway, e.g. Wnt/NOTCH as newly implicated molecular mechanisms in T-PLL pathogenesis for further functional studies. Additionally, novel (KDM6A and KDM6B) and known mutations (ATM, TET2) in chromatin regulatory genes underscore the importance of epigenetic dysregulation in the tumorigenesis of T-PLL.

Disclosures

Parikh: Pharmacyclics: Honoraria, Research Funding. Mustjoki:Ariad: Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Ding:Merck: Research Funding.