Ten-Eleven Translocation 2 (TET2) mutants are frequently associated with hematopoietic malignancies, specifically myeloid and lymphoid cancers. We performed comparative analyses of patient samples across cancer sub-types to characterize TET2 mutations within lymphoid malignancies. Vitamin C is a cofactor for TET enzymes and may restore TET function in specific mutants. This could support vitamin C's role as a supplement in the treatment of hematopoietic malignancies. Patient sample analysis revealed that TET2 mutations in the B cell malignancies are primarily missense mutations and the top missense mutations are located in the N-terminal protein domain, unlike TET2 mutations in myeloid cancers which are primarily located in the C-terminal domain. To investigate these specific TET2 mutations and their functional effect in the N terminus, we prepared a methodology for site-directed mutagenesis of the wild-type TET2 gene. The mutant TET2 proteins of the four most frequent TET2 mutations identified in B lymphoma will be transfected into HEK293T cells for treatment with and without vitamin C to determine effects on TET2 enzymatic activity. We will use Western blot analysis to determine TET2 mutant expressions levels and cellular localization, and dot blot analyses to quantify DNA modifications generated by TET2 enzymatic activity. These studies will allow us to understand if vitamin C has a therapeutic effect on diseased cells with TET2 mutants represented in B Lymphoma.
