Abstract The bone marrow stroma and especially osteoblasts, are critical regulators of the development of hematological myeloid malignancies, actively promoting their development or progression. Such tumorigenic properties of osteoblasts can be pertinent in the context of highly relevant and frequent MDS (and AML) mutations that by themselves are not sufficient to induce disease. One such prominent case are gain-of-function mutations in Isocitrate dehydrogenase 1 and 2 (IDH1/2) drive the synthesis of the oncometabolite 2-hydroxyglutarate (2HG) which induces chromatin abnormalities, including globally altered histone and DNA methylation profiles in MDS (and AML) cells. However, IDH1 and IDH2 mutations are not by themselves sufficient to cause MDS. In addition, IDH1 and IDH2 mutations are detected in expanded clones in aging healthy humans, a phenomenon termed “age related clonal hematopoiesis” that confers increased risk for MDS and AML development. These observations suggest a) that IDH1/2 mutations require additional cooperative signals to become fully transforming; and b) that such signals may accumulate with aging and potentiate the transformation to MDS (or AML). In investigating this possibility in IDH1/2-mutant MDS/AML, we found that D2HG is elevated in the bone marrow plasma of MDS patients carrying IDH mutations as compared to MDS patients without such mutations leading to differential hypermethylation in the stroma of IDH1 and IDH2-mutant patients as compared to IDH1 and IDH2 wild type patients. Integrative analysis of these results with RNAseq data from the same subjects and preliminary in vivo analysis suggests stromal cell candidate genes that affect IDH1/2-related transformations. Importantly, hypermethylation of these molecules increases in osteoblasts with aging and also presents in subjects with IDH1/2 age related clonal hematopoiesis (ARCH). In addition, it correlates with resistance to IDH2 inhibitor treatment in patients. We hypothesize that D2HG uptaken by bone marrow stroma leads to hypermethylation in ARCH subjects and with aging. In turn, signals from hypermethylated genes in osteoblasts, act on HSCs harboring IDH1/2 mutations to promote their expansion and transformation to MDS. We will delineate how epigenetically modified mouse and human stromal cells and osteoblasts promote transformation of IDH2 mutant HSCs to MDS/AML; Determine whether clonal expansion is the mechanism through which epigenetic modifications in aging bone marrow stroma and osteoblasts supports ARCH-related transformation of IDH2-mutant HSCs to MDS/AML-initiating cells; Evaluate the role of stroma hypermethylation in resistance to IDH-inhibitor treatment in patients and in overcoming IDH-inhibitor resistance in mice; Confirm identified and uncover additional stromal components of IDH1/2 transformation. These studies will identify osteoblast-derived signals that may be targeted therapeutically alone or in combination with current IDH1/2 inhibitors to imp...