Diverse Landscape of TET2 Variants in MDS and AML
Authors: Tallis Eran , Benton Christopher B. , Khan Abir , Assi Rita , DiNardo Courtney D. , Hornbaker Marisa J , Major Catherine Kendall , Boddu Prajwal , Kanagal-Shamanna Rashmi , Patel Keyur P. , Daver Naval G. , Takahashi Koichi , Montalban-Bravo Guillermo , Kadia Tapan M. , Konopleva Marina Y. , Andreeff Michael , Kantarjian Hagop M. , Garcia-Manero Guillermo 
Published: October 2019

Abstract
INTRODUCTION
The TET2 (Ten‐Eleven Translocation 2) gene, located at chromosome 4q24.1, belongs to TET family of proteins that possesses the capacity of catalyzing the conversion of 5‐methylcytosine into 5‐hydroxymethylcytosine. It is considered to be a putative tumor suppressor gene during cancer initiation and development. TET2 mutations are extensively studied and reported in a variety of human hematological malignancies including acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), suggesting a crucial role of TET2 in the pathogenesis of blood cancers. The landscape of rare TET2 variants which may be of potential clinical relevance is not fully described. The landscape of TET2 variants from a large cohort of MDS and AML patients was explored.
METHODS
We analyzed data from bone marrow samples of 2634 patients with AML or MDS who presented at MD Anderson between the years 2012-2016. Seven hundred seventy seven patients were found to have TET2 variants that were not classifiable as known mutations. Two hundred sixty-three patients were diagnosed with MDS, 144 with secondary AML transformed from MDS, and 370 with AML without prior history of MDS. The coding regions of TET2 gene was analyzed by next-generation sequencing in the molecular diagnostics laboratory. Non-mutation variants were classified into 2 groups: 1) those reported as germline polymorphisms (GP) in population studies, literature or matched tumor-normal analysis on patients at MDACC; and 2) variants for which a germline versus somatic origin could not be determined unequivocally (variants of uncertain origin, VUO). Very common GP variants (defined as those with a population frequency of over 20% in laboratory sample cohort) were not evaluated. Variants' predicted frequency in the population according to gnomAD Exomes 2.0.2, and classification according to American College of Medical Genetics and Genomics (ACMG) guidelines for interpretation of sequence variants, were assessed using the VarSome platform by Saphetor.
RESULTS
Two-hundred and twenty-eight unique variants were identified. The amino acid substitutions from missense variants were widely distributed throughout TET2. Fifty-four unique variants were classified as GP and several of these recurrent variants were found to be present in our population at a significantly higher frequency (p<0.0001-0.03) than expected based on population sequencing. Within this group, 37 (69%) were missense, 8 (15%) frameshift (fs), 5 (9%) nonsense, and 4 (7%) splice-site. By ACMG classification 10 (27%) of these missense variants were benign, 27 (73%) were uncertain significance (US), and none pathogenic. Benign variants were shared across MDS, MDS to AML, and AML groups. Thirty-seven percent of the variants of US were shared across the groups whereas frameshift and nonsense variants were not shared between groups and were primarily observed in AML patients. One-hundred and seventy-four unique variants classified as VUO were found. Most such variants (110 [63%]) were found in MDS patients, while 32 (29%) were found in the transformed MDS/AML group and 36 (33%) in AML patients. There was a statistically significant higher fraction of VUO in the MDS pts vs AML and AML/MDS pts when compared in the initial 2634 patients (9% vs. 5%, p=0.0002). Among VUO, 105 (60%) were missense, 47 (27%) fs, 12 (7%) nonsense, and 10 (6%) splice-site. In the missense group, none were classified as benign variants, 86 (82%) variants of US, and 19 (18%) pathogenic variants.
CONCLUSIONS
The landscape of rare TET2 variants in a large group of MDS and AML patients was frequent, diverse, and included both GP and VUO. GP variants were more likely benign by ACMG classification and found in both MDS and AML patients. A large number of singleton VUO were found among MDS patients, and many were predicted to be pathogenic or had uncertain significance. VUO's also contained many fs and nonsense variants. These result support the idea that rare pathogenic variants in TET2 are currently underrecognized as contributory, although they are likely to help drive leukemogenesis. Greater understanding of the importance of individual TET2 variants and additional classification metrics are needed.

Disclosures
DiNardo: Karyopharm: Honoraria; Abbvie: Honoraria; Agios: Consultancy; Medimmune: Honoraria; Celgene: Honoraria; Bayer: Honoraria. Daver:Karyopharm: Research Funding; Incyte: Consultancy; Novartis: Consultancy; ImmunoGen: Consultancy; Sunesis: Consultancy; Pfizer: Consultancy; Daiichi-Sankyo: Research Funding; Kiromic: Research Funding; Alexion: Consultancy; Otsuka: Consultancy; Sunesis: Research Funding; Incyte: Research Funding; ARIAD: Research Funding; BMS: Research Funding; Karyopharm: Consultancy; Novartis: Research Funding; Pfizer: Research Funding. Kadia:Celgene: Research Funding; Takeda: Consultancy; BMS: Research Funding; Abbvie: Consultancy; Novartis: Consultancy; Novartis: Consultancy; Amgen: Consultancy, Research Funding; Jazz: Consultancy, Research Funding; Abbvie: Consultancy; Pfizer: Consultancy, Research Funding; Takeda: Consultancy; Amgen: Consultancy, Research Funding; BMS: Research Funding; Pfizer: Consultancy, Research Funding; Jazz: Consultancy, Research Funding; Celgene: Research Funding. Konopleva:Stemline Therapeutics: Research Funding; abbvie: Research Funding; cellectis: Research Funding; Immunogen: Research Funding. Andreeff:Amgen: Consultancy, Research Funding; Astra Zeneca: Research Funding; Jazz Pharma: Consultancy; Oncolyze: Equity Ownership; Aptose: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Oncoceutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; SentiBio: Equity Ownership; United Therapeutics: Patents & Royalties: GD2 inhibition in breast cancer ; Reata: Equity Ownership; Celgene: Consultancy; Eutropics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Consultancy, Patents & Royalties: MDM2 inhibitor activity patent, Research Funding.