Abstract
Ten-eleven translocation 2 (TET2) encodes a DNA demethylation enzyme, and it is one of the most frequently mutated genes in myeloid neoplasms and clonal hematopoiesis of indeterminate potential (CHIP). The large size of the TET2 gene, spanning approximately 134 kilobases, makes it particularly susceptible to truncating mutations. Although TET2 mutations are highly heterogeneous, the majority result in the loss of enzymatic function, which is found across a broad spectrum of myeloid disorders.
A characteristic clinical feature of TET2-mutant myeloid neoplasms, particularly chronic myelomonocytic leukemia (CMML), is persistent monocytosis. TET2 mutations also correlate with a broader phenotype of myeloid lineage bias and clonal expansion, including increased granulocyte-monocyte progenitor (GMPs) and monocyte output in the setting of CHIP in humans. Using a Tet2-/- bone marrow transplantation (BMT) model, we found that Gasdermin D (Gsdmd), a key mediator of the inflammatory responses in the bone marrow microenvironment, is essential for the development of myelodysplastic syndrome (MDS)/CMML-like phenotypes in mice, including monocytosis and anemia. Ablation of Gsdmd in the bone marrow microenvironment prevented these phenotypes, demonstrating that Tet2 mutation drives disease development through a cell-non-autonomous mechanism.
Interestingly, we also observed a marked expansion of CD4+ T cells in the bone marrow of Tet2-/- BMT mice, which was partially rescued by the Gsdmd null bone marrow microenvironment. Single-cell RNA sequencing revealed that the most prominently expanded CD4+ T cell populations exhibited transcriptional signatures of regulatory T (Treg) cells and T follicular helper (Tfh) cells, a finding further validated by flow cytometry analysis. Using a multiplex cytokine assay, we identified a distinct cluster of cytokines elevated in the bone marrow fluid of Tet2-/- BMT mice, including IL-6, IL-1β, IL-12p70, and CD40L. Notably, CD4+ T cells isolated from eight-month-old Tet2-/- mice with CMML-like disease were able to drive monocytic expansion and significantly accelerate disease progression, even in the Gsdmd-null microenvironment. These findings suggest a potential crosstalk between monocytes/macrophages and CD4+ T cells in the Tet2-/- bone marrow microenvironment. Specifically, macrophage-derived IL-12p70 promotes CD4+ T cell expansion, while CD4+ T cell–derived CD40L, in turn, facilitates monocytic expansion.
The insights gained from the Tet2-/- BMT mouse model are highly clinically relevant. To further investigate CD4+ T cell expansion in humans, we examined a cohort of patients diagnosed with myeloid disorders, including clonal cytopenia of undetermined significance (CCUS), MDS, myeloproliferative neoplasms (MPN), and CMML, all of whom harbored isolated somatic TET2 mutations (n = 9). Treatment-naïve lymphoma patients without bone marrow involvement served as the normal control group (n = 10). The bone marrow CD4/CD8 ratio was used to assess the expansion of CD4+ T cells. The TET2-mutant group exhibited a significantly higher CD4/CD8 ratio compared to the control group. To determine whether CD4+ T cell expansion is specific to TET2 mutations, we analyzed a separate cohort of patients with myeloid disorders and isolated somatic DNMT3A mutations (n = 8). In this group, there was no significant difference in the CD4/CD8 ratio compared to controls, suggesting that CD4+ T cell expansion may be exclusive to TET2-mutant cases. Additionally, review of bone marrow core biopsy from the TET2-mutation group revealed that a high CD4/CD8 ratio was associated with the presence of tertiary lymphoid structures in the bone marrow. These structures were characterized by a core of Tfh cells surrounded by B lymphocytes. This observation in human samples is consistent with our findings in the Tet2-/- BMT mouse model, where a distinct population of Tfh-like cells was also identified.
Together, these findings reveal a previously unrecognized connection between TET2 mutations and the expansion of bone marrow CD4+ T cells, triggered by the inflammatory bone marrow microenvironment, in myeloid disorders. Our results suggest that TET2-driven immune remodeling may contribute to disease pathogenesis by fostering aberrant interactions between the myeloid and lymphoid compartments.
