Loss of epigenetic regulator TET2 and oncogenic KIT regulate myeloid cell transformation via PI3K pathway
Lakshmi Reddy Palam, Raghuveer Singh Mali, Baskar Ramdas, Sridhar Nonavinkere Srivatsan, Valeria Visconte, Ramon V. Tiu, Bart Vanhaesebroeck, Axel Roers, Alexander Gerbaulet, Mingjiang Xu, Sarath Chandra Janga, Clifford M. Takemoto, Sophie Paczesny, Reuben Kapur
Lakshmi Reddy Palam, Raghuveer Singh Mali, Baskar Ramdas, Sridhar Nonavinkere Srivatsan, Valeria Visconte, Ramon V. Tiu, Bart Vanhaesebroeck, Axel Roers, Alexander Gerbaulet, Mingjiang Xu, Sarath Chandra Janga, Clifford M. Takemoto, Sophie Paczesny, Reuben Kapur
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Research Article Hematology

Loss of epigenetic regulator TET2 and oncogenic KIT regulate myeloid cell transformation via PI3K pathway

Abstract

Mutations in KIT and TET2 are associated with myeloid malignancies. We show that loss of TET2-induced PI3K activation and -increased proliferation is rescued by targeting the p110α/δ subunits of PI3K. RNA-Seq revealed a hyperactive c-Myc signature in Tet2–/– cells, which is normalized by inhibiting PI3K signaling. Loss of TET2 impairs the maturation of myeloid lineage–derived mast cells by dysregulating the expression of Mitf and Cebpa, which is restored by low-dose ascorbic acid and 5-azacytidine. Utilizing a mouse model in which the loss of TET2 precedes the expression of oncogenic Kit, similar to the human disease, results in the development of a non–mast cell lineage neoplasm (AHNMD), which is responsive to PI3K inhibition. Thus, therapeutic approaches involving hypomethylating agents, ascorbic acid, and isoform-specific PI3K inhibitors are likely to be useful for treating patients with TET2 and KIT mutations.

Authors

Lakshmi Reddy Palam, Raghuveer Singh Mali, Baskar Ramdas, Sridhar Nonavinkere Srivatsan, Valeria Visconte, Ramon V. Tiu, Bart Vanhaesebroeck, Axel Roers, Alexander Gerbaulet, Mingjiang Xu, Sarath Chandra Janga, Clifford M. Takemoto, Sophie Paczesny, Reuben Kapur

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Figure 8

TET2 loss in cooperation with oncogenic KIT induced myeloproliferative neoplasm.

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TET2 loss in cooperation with oncogenic KIT induced myeloproliferative n...
(A) WT, Tet2+/–, and Tet2+/–:Kit+/D814V mice were intraperitoneally injected with poly(I:C), and mice were analyzed after 12–16 weeks of treatment. Flow cytometric quantification on BM cells was performed by staining the cells with KIT, Sca-1, and lineage-specific antibodies. The percentage of cells negatively selected for lineage markers, KIT positivity, and Sca-1 positivity (LSK) is shown above inset boxes. Peripheral blood and BM cells were also analyzed by flow cytometry using Mac1, Gr1, and KIT antibodies, and the percentage of cells positive for Mac1 and Gr1 or cells positive for Mac1 and KIT is shown in dot blots. Spleen images of indicated genotypes are shown. Experiments were performed more than 3 times, and data from a representative experiment are shown. (B) BM-derived mast cells (BMMCs) were generated from WT, Tet2+/–, and Tet2+/–:Kit+/D814V mice described in A. Cells were harvested and stained with antibodies that recognize KIT and IgE receptor after the indicated weeks of culture, followed by flow cytometry analysis. The percentage of BMMCs double positive for KIT and IgE receptor expression are shown in dot blots. Representative of 3 independent experiments. (C) BMMCs as cultured in B were harvested, and Tet2, Kit, Cebpa, Mitf, and PU.1 mRNA levels, relative to β-actin mRNA levels, were measured using QRT-PCR. Mean ± SD, * P < 0.05. (D) BMMCs from mice with indicated genotypes were starved for 6 hours and cultured in the presence or absence of IL-3 or SCF. After 48 hours, proliferation was evaluated by [3H] thymidine incorporation. Counts per minute (CPM) are shown. n = 3, mean ± SEM, *P < 0.05. One-way ANOVA test used for statistical analysis (C and D).