Design, synthesis and biological activity of 4-(4-benzyloxy)phenoxypiperidines as selective and reversible LSD1 inhibitors
Graphical abstract
Introduction
Epigenetic post-transcriptional modifications on histones, including methylation, acetylation and phosphorylation, regulate gene activation and repression [1]. Lysine specific demethylase 1 (LSD1), the first histones demethylase discovered in 2004, is a highly conserved flavin adenine dinucleotide (FAD) dependent oxidative enzyme, which could catalyze mono- and dimethyl histone 3 lysine 4 (H3K4me1, H3K4me2) and histone 3 lysine 9 (H3K9me1 and H3K9me2) demethylation [2], [3]. In addition, LSD1 could also demethylate nonhistones substrates including p53, DNA methyltransferase 1 (DNMT1), signal transducer and activator of transcription 3 (STAT3), E2F transcription factor 1 and regulates the downstream cellular functions [4], [5], [6], [7], [8]. Overexpression of LSD1 is observed in acute myeloid leukemia (AML), ER-negative breast cancer, prostate cancer, glioblastoma, degenerative diseases [9], [10], [11], [12]. Moreover, downregulation of LSD1 expression by inhibition with small molecules can suppress the proliferation and metastasis in cancer cells, as well as inducing differentiation and apoptosis of gliama stem cells and selectively targeting cancer cells with pluripotent stem cell properties [13], [14]. Therefore, LSD1 is an attractive anti-cancer drug target.
To date, a variety of LSD1 inhibitors with diverse chemotypes have been identified (Fig. 1) [9], [15]. As a member of monoamine oxidase (MAO) family, LSD1 shares highly similarity with the homologous MAOs in sequence prompted the investigation of anti-MAOs drugs as potential LSD1 inhibitors. Tranylcypromine (TCPA, A), a widely used MAO inhibitor with antidepressant properties, was reported to be a mechanised-based irreversible and poor selective LSD1 inhibitor at clinical doses. The mechanism for TCPA is that binds directly to LSD1’s catalytic site via an irreversible covalent interaction [16]. Since then, a series of TCPA-based inhibitors with high selectivity and potency have been emerged. ORY-1001 (C), developed by Oryzon Genomics, is in Phase I/IIa for the treatment of AML [17]. GSK2879552 (B) with high growth inhibitory is in Phase I trial to treat relapsed/refractory small cell lung cancer (SCLC) [18], [19]. Based on the TCPA scaffold, some fluorinated and spirocyclic derivatives have been recently reported with potent inhibitory activities [20], [21], [22]. Besides irreversible inhibitors, reversible LSD1 inhibitors have been attracted plenty of attention. D and E selectively inhibit LSD1 at micromolar level and these kinds of inhibitors deserving further structural modification [23], [24], [25]. GSK-354 (F) is a potent reversible LSD1 inhibitors with Ki values as low as 29 nM [26]. SP-2509 (G) is a potent, reversible and specific inhibitors with an IC50 value of 13 nM in vitro [27]. Optimization of SP-2509 via conformation constrain strategy afforded a series of novel (E)-N′-(2,3-dihydro-1H-inden-1-ylidene) benzohydrazides and the best compound displayed its potency with IC50 value of 1.4 nM [28]. Other reversible LSD1 inhibitors have been recently reported [29], [30]. Recently, the structure of thieno-[3,2-b]pyrrole-5-carboxamides (H) as novel LSD1 reversible inhibitors with the IC50 value of 7.8 nM which displays a remarkable anticlonogenic cell growth effect on MLL-AF9 human leukemia cells [31], [32].
Taken the co-crystal structure of LSD1 and H into consideration, a series of derivatives of thieno-[3,2-b]pyrrole-5-carboxamides with other aromatic rings were designed and synthesized to explore more potent LSD1 inhibitors (Fig. 2). Importantly, H was investigated in hematological diseases not solid tumors. Therefore, we wanted to obtain a series of its analogs (10a-10g) for the treatment of solid tumors. Compound 10d was identified as a potent, selective, and reversible LSD1 inhibitors at single-digit micromolar level and could inhibit the migration of A549 cell line, which deserves the further structural optimization.
Section snippets
Chemistry
Synthetic route for the target compounds 11a-11g was depicted in Scheme 1. 2-Methyl-6-nitrobenzoic acid as the starting material was methylated with iodomethane to obtain methyl-2-methyl-6-nitrobenzoate (1a). Bromination of 1a with NBS and AIBN in carbon tetrachloride gave 2a. 2a was reacted with sodium hydride to obtain methyl 2-(methoxymethyl)-6-nitrobenzoate (3a). Subsequent reduction of methyl ester gave 4a, which was transferred into its bromo-derivative 5a. 5a was reacted with
Conclusion
In summary, we designed and synthesized a series of 4-(4-benzyloxy)phenoxypiperidines as reversible LSD1 inhibitors. Compound 10d displayed a highly selective inhibitory activity against LSD1 with an IC50 value of 4 μM. Moreover, wound healing assay indicated that 10d owns a positive effect for the migration of HCT-116 colon cancer cells and A549 lung cancer cells. Thus, 10d deserves further investigation as a hit-to-lead for the treatment of LSD1 associated tumors.
Chemistry
Regents and solvents were purchased from Sinopharm Chemical Reagent Co. Ltd. (Shanghai, China). Thin-layer chromatography (TLC) was carried out on silica gel GF254 plates (Qingdao Haiyang Chemical Plant, Qingdao, Shandong, China) and the spots were visualized using a UV lamp (λ = 254 nm). The purification of products were performed on silica gel columns (90–150 μm; Qingdao Marine Chemical Inc., Qingdao, Shandong, China) by flash chromatography. Melting points were measured on an XT-4
Notes
F. L. is a shareholder of Constellation Pharmaceuticals Inc. as well as a consultant of Active Motif.
Acknowledgments
X. Z. thanks the Natural Science Foundation of Jiangsu Province (BK20161458), “Six Talent Peaks” Project of Jiangsu Province (2016-YY-042) and Postgraduate Research & Practice Innovation Program of Jiangsu Province (KYCX17_0721). F. L. thanks the financial support by China “Thousand Youth Talents” (KHH1340001) and NSFC grant (91419306).
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These authors contributed equally to this work.