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.
ARTICLE ABSTRACT
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
