American Association for Cancer Research
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Supplementary Figures 1 - 7 from Mechanism, Consequences, and Therapeutic Targeting of Abnormal IL15 Signaling in Cutaneous T-cell Lymphoma

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journal contribution
posted on 2023-04-03, 21:03 authored by Anjali Mishra, Krista La Perle, Sonya Kwiatkowski, Laura A. Sullivan, Gregory H. Sams, Jessica Johns, Douglas P. Curphey, Jing Wen, Kathleen McConnell, Jun Qi, Henry Wong, Giandomenico Russo, Jianying Zhang, Guido Marcucci, James E. Bradner, Pierluigi Porcu, Michael A. Caligiuri

Supplementary Figures 1 - 7. Supplementary Figure 1. Immunoblot analysis of IL-15 in normal donor CD4+ T-cells, SS patient CD4+ T-cells and patient derived cell lines. Supplementary Figure 2. Representative flow cytometric histogram is shown for CD3 expression in mononuclear cells from skin of WT and IL-15 tg mice. Supplementary Figure 3. Immunoblot analysis of HDAC1 in normal donor CD4+ T-cells, SS patient CD4+ T-cells and patient derived cell line, Hut78. Protein lysates were probed for expression of HDAC1; β-Actin was used as housekeeping control. Supplementary Figure 4. Immunoblot analysis for expression of p21 protein in normal donor CD4+ cells following ten days of incubation in IL-15 compared to unstimulated normal donor CD4+ T-cells. Supplementary Figure 5. Pie chart showing the distribution of HDAC1 occupancy in normal donor CD4+ T-cells (upper left) compared to patient CD4+ T-cells (upper right) and normal donor CD4+ T-cells stimulated with 100ng/ml IL-15 for 24 hours (lower left). Supplementary Figure 6. High magnification and (B) low magnification histological analysis of IL-15 Tg skin. Supplementary Figure 7. The CTCL mouse disease severity scoring system is on a 0 to 100 scale, with 100 being the highest (most severe) score.



Cutaneous T-cell lymphoma (CTCL) is the most common type of primary cutaneous lymphoma. Here, we report that patients with CTCL show increased IL15 in a clinical stage–dependent manner. Mechanistically, we show that ZEB1 is a transcriptional repressor of IL15 in T cells and that hypermethylation of the ZEB1 binding region within the IL15 promoter, as seen in patients with CTCL, prevents ZEB1 binding and causes increased transcription of IL15. Using a transgenic mouse model of IL15, we provide evidence that overexpression of IL15 induces a spontaneous CTCL that mimics the human neoplasm. Excessive autocrine production of IL15 in T cells inhibits an HDAC1-mediated negative autoregulatory loop, resulting in the upregulation of HDAC1 and HDAC6 and transcriptional induction of the onco-miR-21. Interruption of IL15 downstream signaling with isotype-specific HDAC inhibitors halts (HDAC1) or significantly delays (HDAC6) the progression of CTCL in vivo and provides preclinical evidence supporting a hierarchical model of oncogenic signaling in CTCL.Significance: To date, CTCL pathogenesis remains unknown, and there are no curative therapies. Our findings not only demonstrate a critical role for IL15-mediated inflammation in cutaneous T-cell lymphomagenesis, but also uncover a new oncogenic regulatory loop in CTCL involving IL15, HDAC1, HDAC6, and miR-21 that shows differential sensitivity to isotype-specific HDAC inhibitors. Cancer Discov; 6(9); 986–1005. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 932