Region-specific alteration of histone modification by LSD1 inhibitor conjugated with pyrrole-imidazole polyamide

Epigenome regulates gene expression to determine cell fate, and accumulation of epigenomic aberrations leads to diseases, including cancer. NCD38 inhibits lysine-specific demethylase-1 (LSD1), a histone demethylase targeting H3K4me1 and H3K4me2, but not H3K4me3. In this study, we conjugated NCD38 with a potent small molecule called pyrrole (Py) imidazole (Im) polyamide, to analyze whether targets of the inhibitor could be regulated in a sequence-specific manner. We synthesized two conjugates using β-Ala (β) as a linker, i.e., NCD38-β-β-Py-Py-Py-Py (NCD38-β(2)P(4)) recognizing WWWWWW sequence, and NCD38-β-β-Py-Im-Py-Py (NCD38-β(2)PIPP) recognizing WWCGWW sequence. When RKO cells were treated with NCD38, H3K4me2 levels increased in 103 regions with significant activation of nearby genes (P = 0.03), whereas H3K4me3 levels were not obviously increased. H3K27ac levels were also increased in 458 regions with significant activation of nearby genes (P = 3 × 10(−10)), and these activated regions frequently included GC-rich sequences, but less frequently included AT-rich sequences (P < 1 × 10(−15)) or WWCGWW sequences (P = 2 × 10(−13)). When treated with NCD38-β(2)P(4), 234 regions showed increased H3K27ac levels with significant activation of nearby genes (P = 2 × 10(−11)), including significantly fewer GC-rich sequences (P < 1 × 10(−15)) and significantly more AT-rich sequences (P < 1 × 10(−15)) compared with NCD38 treatment. When treated with NCD38-β(2)PIPP, 82 regions showed increased H3K27ac levels, including significantly fewer GC-rich sequences (P = 1 × 10(−11)) and fewer AT-rich sequences (P = 0.005), but significantly more WWCGWW sequences (P = 0.0001) compared with NCD38 treatment. These indicated that target regions of epigenomic inhibitors could be modified in a sequence-specific manner and that conjugation of Py-Im polyamides may be useful for this purpose.