Selective disruption of RORγt-CBFβ interaction by IMU-935 prevents RORγt-dependent Th17 autoimmunity but not thymocyte development
Hongmin Wu, Xiancai Zhong, Ning Ma, Zhiheng He, Guanpeng Wang, Geming Lu, Yate-Ching Yuan, Wencan Zhang, Yun Shi, Nagarajan Vaidehi, Evelyn Peelen, Tanja Wulff, Christian Gege, Hella Kohlhof, Daniel Vitt, Yousang Gwack, Ichiro Taniuchi, Hai-Hui Xue, Zuoming Sun
Hongmin Wu, Xiancai Zhong, Ning Ma, Zhiheng He, Guanpeng Wang, Geming Lu, Yate-Ching Yuan, Wencan Zhang, Yun Shi, Nagarajan Vaidehi, Evelyn Peelen, Tanja Wulff, Christian Gege, Hella Kohlhof, Daniel Vitt, Yousang Gwack, Ichiro Taniuchi, Hai-Hui Xue, Zuoming Sun
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Research Article Autoimmunity Immunology

Selective disruption of RORγt-CBFβ interaction by IMU-935 prevents RORγt-dependent Th17 autoimmunity but not thymocyte development

Abstract

RORγt is a key transcription factor regulating both Th17 differentiation and thymocyte development. Although Th17 cells drive autoimmune diseases, inhibiting RORγt to treat autoimmunity also disrupts thymocyte development and can cause lethal thymic lymphoma. We identified a previously unreported RORγt cofactor, CBFβ, and a highly selective RORγt inhibitor, IMU-935, that preferentially disrupt the RORγt-CBFβ interaction in Th17 cells but not thymocytes. This interaction is essential for RORγt function; mice with a RORγt mutant unable to bind CBFβ had impaired Th17 differentiation, were resistant to experimental autoimmune encephalomyelitis (EAE), and had defective thymocyte development. IMU-935 inhibited Th17 differentiation and reduced EAE severity without affecting thymocyte development by selectively targeting the RORγt-CBFβ interaction in Th17 cells but not in thymocytes. This differential effect arose because different concentrations of IMU-935 were required to disrupt the interaction in Th17 cells versus thymocytes, due to varying levels of RUNX1 that compete with RORγt for CBFβ binding. This study reveals an unreported mechanism for RORγt regulation and a selective RORγt inhibitor that prevents Th17-driven autoimmunity without the risk of lethal lymphoma from thymocyte disruption.

Authors

Hongmin Wu, Xiancai Zhong, Ning Ma, Zhiheng He, Guanpeng Wang, Geming Lu, Yate-Ching Yuan, Wencan Zhang, Yun Shi, Nagarajan Vaidehi, Evelyn Peelen, Tanja Wulff, Christian Gege, Hella Kohlhof, Daniel Vitt, Yousang Gwack, Ichiro Taniuchi, Hai-Hui Xue, Zuoming Sun

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Figure 5

IMU-935 does not affect genes critical for thymocyte survival.

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IMU-935 does not affect genes critical for thymocyte survival. (A) Venn ...
(A) Venn diagram displaying the number of DEGs (1.5-fold, up or down; P < 0.05) identified by RNA-Seq assays in WT versus RORγt–/– thymocytes and in WT thymocytes treated with IMU-935 (1 μM) or not. (B and C) Volcano plots displaying the global gene expression. The horizontal dotted line marks P = 0.05, the vertical dotted lines mark fold change of ±1.5. (D) Heatmap showing antiapoptotic, apoptotic, and cell cycle gene expression in RORγt–/– or IMU-935–nontreated (WT-vehicle control) or treated (WT–IMU-935) thymocytes. Gene expression was normalized by total counts of each gene. (E) Heatmap depicting changes in the activity of upstream regulators predicted by IPA. Activation z score indicates increased (orange; z > 0) or decreased (blue; z < 0) activity. All z score values are labeled within the corresponding cell. Genes shown are known to regulate apoptosis (Mycn, E2f1, Esr1), survival (Nfkbia, TP53), cell cycle (Gli1, Cdkn2a), oxidative stress (Nfe2l2), and thymocyte development (Ctnnb1, Lef1). (F) Heatmap showing the activity of the pathways determined by IPA. Pathway activity was obtained by comparing WT and RORγt–/– thymocytes (left column) and IMU-935–nontreated versus -treated WT thymocytes (right column). N/A indicates pathway status is not predictable due to the small number of genes affected by IMU-935 treatment in thymocyte RNA-Seq. (G) Genome-wide RORγt-DNA binding signal intensity determined by ChIP-Seq assays near the TSS in RORγt–/– or WT (RORγt+) or WT–IMU-935 thymocytes. (H) RORγt binding peaks at the Bcl2l1 locus. The results are the representative of overlays from 2 separate experiments. Veh, vehicle.