Identification of selective and reversible LSD1 inhibitors with anti-metastasis activity by high-throughput docking

The overexpression of lysine specific demethylase 1 (LSD1) has been reported in various human tumors. There is increasing interest in targeting LSD1 with small molecules for cancer treatment. A released structure of an LSD1 kinase domain in complex with FAD was used to set up a low-cost high-throughput docking protocol for quick identification of LSD1 inhibitors. The most promising hit L05 was confirmed to be a potent, selective and reversible LSD1 inhibitor and displayed marked inhibition of colorectal cells migration without significant cytotoxicity.     

Natural withanolide-based lysine-specific demethylase 1 inhibitors for antitumor metastasis activity

Lysine specific demethylase 1 (LSD1), which is overexpressed in several human cancers and acts as a demethylase of histone 3, lysine 4 and lysine 9, has become an attractive therapeutic target for cancer therapy. Based on our previous systematic studies, withanolides are important secondary metabolites mostly from the Solanaceae family of plants, which are crucial agents for cancer treatment. Here, withanolides were characterized as LSD1 inhibitors, especially withaferin A, with an IC₅₀ value of 3.04 μM. In vitro bioactivity assays and virtual molecular docking indicated that withaferin A could inhibit MDA-MB-231 cell migration by inhibiting intracellular LSD1 activity. These findings provide a new withanolide-based natural molecular skeleton for LSD1 inhibitors with potential antitumor activity.     

Discovery of tranylcypromine analogs with an acylhydrazone substituent as LSD1 inactivators: Design,synthesis and their biological evaluation

Lysine specific demethylase 1 (LSD1), the first identified histone demethylase, plays an important role in epigenetic regulation of gene activation and repression, has been reported to be up-regulated and involved in numbers of solid malignant tumors. In this study, we identified a series of phenylalanyl hydrazones based LSD1 inhibitors, and the most potent one, compound 4q, can inactivate LSD1 with IC₅₀?=?91.83?nM. In cellular level, compound 4q can induce the accumulation of CD86 as well as H3K4me2, and inhibit gastric cancer cell proliferation by inactivating LSD1. Our findings indicated that compound 4q may serve as a potential leading compound to target LSD1 overexpressed gastric cancer.     

Structure-based design and discovery of potent and selective KDM5 inhibitors

Histone lysine demethylases (KDMs) play a key role in epigenetic regulation and KDM5A and KDM5B have been identified as potential anti-cancer drug targets. Using structural information from known KDM4 and KDM5 inhibitors, a potent series of pyrazolylpyridines was designed. Structure-activity relationship (SAR) exploration resulted in the identification of compound 33, an orally available, potent inhibitor of KDM5A/5B with promising selectivity. Potent cellular inhibition as measured by levels of tri-methylated H3K4 was demonstrated with compound 33 in the breast cancer cell line ZR-75-1.     

Design,synthesis and biological activity of 4-(4-benzyloxy)phenoxypiperidines as selective and reversible LSD1 inhibitors

Lysine specific demethylase 1 (LSD1) plays a vital role in epigenetic regulation of gene activation and repression in several human cancers and is recognized as a promising antitumor therapeutic target. In this paper, a series of 4-(4-benzyloxy)phenoxypiperidines were synthesized and evaluated. Among the tested compounds, compound 10d exhibited the potent and reversible inhibitory activity against LSD1 in vitro (IC₅₀ = 4 μM). Molecular docking was conducted to predict its binding mode. Furthermore, 10d displayed it could inhibit migration of HCT-116 colon cancer cells and A549 lung cancer cells. Taken together, 10d deserves further investigation as a hit-to-lead for the treatment of LSD1 associated tumors.     

Development of 5-hydroxypyrazole derivatives as reversible inhibitors of lysine specific demethylase 1

A series of reversible inhibitors of lysine specific demethylase 1 (LSD1) with a 5-hydroxypyrazole scaffold have been developed from compound 7, which was identified from the patent literature. Surface plasmon resonance (SPR) and biochemical analysis showed it to be a reversible LSD1 inhibitor with an IC₅₀ value of 0.23 µM. Optimisation of this compound by rational design afforded compounds with K_d values of <10 nM. In human THP-1 cells, these compounds were found to upregulate the expression of the surrogate cellular biomarker CD86. Compound 11p was found to have moderate oral bioavailability in mice suggesting its potential for use as an in vivo tool compound.     

Design,synthesis and biological evaluation of curcumin analogues as novel LSD1 inhibitors

Histone lysine-specific demethylase 1 (LSD1) was the first discovered histone demethylase. Inactivating LSD1 or downregulating its expression inhibits cancer-cell development, and thus, it is an attractive molecular target for the development of novel cancer therapeutics. In this study, we worked on the structural optimization of natural products and identified 30 novel LSD1 inhibitors. Utilizing a structure-based drug design strategy, we designed and synthesized a series of curcumin analogues that were shown to be potent LSD1 inhibitors in the enzyme assay. Compound WB07 displayed the most potent LSD1 inhibitory activity, with an IC₅₀ value of 0.8 μM. Moreover, WA20 showed an anticlonogenic effect on A549 cells with an IC₅₀ value of 4.4 μM. Molecular docking simulations were also carried out, and the results indicated that the inhibitors bound to the protein active site located around the key residues of Asp555 and Asp556. These findings suggested that compounds WA20 and WB07 are the first curcumin analogue-based LSD1 inhibitors with remarkable A549 suppressive activity, providing a novel scaffold for the development of LSD1 inhibitors.     

Design,synthesis and evaluation of γ-turn mimetics as LSD1-selective inhibitors

Lysine-specific demethylase 1 (LSD1) is an attractive molecular target for cancer therapy. We have previously reported potent LSD1-selective inhibitors (i.e., NCD18, NCD38, and their analogs) consisting of trans-2-phenylcyclopropylamine (PCPA) or trans-2-arylcyclopropylamine (ACPA) and a lysine moiety that could form a γ-turn structure in the active site of LSD1. Herein we report the design, synthesis and evaluation of γ-turn mimetic compounds for further improvement of LSD1 inhibitory activity and anticancer activity. Among a series of γ-turn mimetic compounds synthesized by a Mitsunobu-reaction-based amination strategy, we identified 1n as a potent and selective LSD1 inhibitor. Compound 1n induced cell cycle arrest and apoptosis through histone methylation in human lung cancer cells. The γ-turn mimetics approach should offer new insights into drug design for LSD1-selective inhibitors.     

Discovery of orally active chalcones as histone lysine specific demethylase 1 inhibitors for the treatment of leukaemia

Histone lysine specific demethylase 1 (LSD1) has emerged as an attractive molecule target for the discovery of potently anticancer drugs to treat leukaemia. In this study, a series of novel chalcone derivatives were designed, synthesised and evaluated for their inhibitory activities against LSD1 in vitro. Among all these compounds, D6 displayed the best LSD1 inhibitory activity with an IC₅₀ value of 0.14 μM. In the cellular level, compound D6 can induce the accumulation of H3K9me1/2 and inhibit cell proliferation by inactivating LSD1. It exhibited the potent antiproliferative activity with IC₅₀ values of 1.10 μM, 3.64 μM, 3.85 μM, 1.87 μM, 0.87 μM and 2.73 μM against HAL-01, KE-37, P30-OHK, SUP-B15, MOLT-4 and LC4-1 cells, respectively. Importantly, compound D6 significantly suppressed MOLT-4 xenograft tumour growth in vivo, indicating its great potential as an orally bioavailable candidate for leukaemia therapy.     

Design,synthesis and biological evaluation of benzimidazole derivatives as novel human Pin1 inhibitors

In this work, a series of novel benzimidazole derivatives were designed and synthesized as Pin1 inhibitors. Protease-coupled assay was used to investigate the Pin1 inhibitory potency of all synthesized compounds. Thirteen of them showed preferable Pin1 inhibitory effects with IC₅₀ values lower than 5 μM, and 12a, 15b, 15d and 16c exhibited the most promising Pin1 inhibitory activity at low micromolar level (0.33–1.00 μM) than the positive control compound Juglone. Flow cytometry results showed that treating PC-3 cells with 16c caused slight cycle arrest in a concentration-dependent manner. The structure-activity relationships of R¹, R², R³ and linker of the benzimidazole derivatives were analyzed in detail, which would help further exploration of new Pin1 inhibitors.     

Histone demethylation catalysed by LSD1 is a flavin-dependent oxidative process

Lysine-specific histone demethylase 1 (LSD1) is a very recently discovered enzyme which specifically removes methyl groups from Lys4 of histone 3. We have addressed the functional properties of the protein demonstrating that histone demethylation involves the flavin-catalysed oxidation of the methylated lysine. The nature of the substrate that acts as the electron acceptor required to complete the catalytic cycle was investigated. LSD1 converts oxygen to hydrogen peroxide although this reactivity is not as pronounced as that of other flavin-dependent oxidases. Our findings raise the possibility that in vivo LSD1 might not necessarily function as an oxidase, but it might use alternative electron acceptors.     

Histone H3 peptides incorporating modified lysine residues as lysine-specific demethylase 1 inhibitors

Lysine-specific demethylase 1 (LSD1) is a flavin-dependent enzyme that removes methyl groups from mono- or dimethylated lysine residues at the fourth position of histone H3. We have previously reported several histone H3 peptides containing an LSD1 inactivator motif at Lys-4. In this study, histone H3 peptides having a trans-2-phenylcyclopropylamine (PCPA), a 2,5-dihydro-1H-pyrrole, and a 1,2,3,6-tetrahydropyridine moiety at Lys-4 were prepared along with related compounds possessing a shorter side chain at the fourth position. Enzymatic assays showed that PCPA peptides containing a longer side chain, which can react with FAD in the active site, are potent LSD1-selective inhibitors.     

Design,synthesis and biological activity of 3-oxoamino-benzenesulfonamides as selective and reversible LSD1 inhibitors

Lysine specific demethylase 1 (LSD1) is a flavin-dependent amine oxidase that selectively removes one or two methyl groups from H3 at Lys4 and is recognized as a promising therapeutic target for cancer and other diseases. Here, a series of 3-oxoamino-benzenesulfonamides were synthesized and evaluated for their inhibitory activity against LSD1. Compounds 7b and 7h showed the most potent inhibition with the IC₅₀ values of 9.5 and 6.9 μM, respectively. Furthermore, the LSD1 inhibition of 7b and 7h were reversible and selective. Docking study presented the possible binding mode between 7b, 7h and the LSD1 active site. Taken together, 3-oxoamino-benzenesulfonamides may represent a new class of reversible LSD1 inhibitors and 7b and 7h were two hit compounds deserved further structural optimization.     

Development and evaluation of 4-(pyrrolidin-3-yl)benzonitrile derivatives as inhibitors of lysine specific demethylase 1

As part of our ongoing efforts to develop reversible inhibitors of LSD1, we identified a series of 4-(pyrrolidin-3-yl)benzonitrile derivatives that act as successful scaffold-hops of the literature inhibitor GSK-690. The most active compound, 21g, demonstrated a K_d value of 22 nM and a biochemical IC₅₀ of 57 nM. In addition, this compound displayed improved selectivity over the hERG ion channel compared to GSK-690, and no activity against the related enzymes MAO-A and B. In human THP-1 acute myeloid leukaemia cells, 21g was found to increase the expression of the surrogate cellular biomarker CD86. This work further demonstrates the versatility of scaffold-hopping as a method to develop structurally diverse, potent inhibitors of LSD1.     

Discovery of vanadium complexes bearing tridentate schiff base ligands as novel LSD1 inhibitors

Lysine specific demethylase (LSD1) plays a pivotal role in epigenetic modulation of gene expression. Abberrant expression of LSD1 was associated with the progress and oncogenesis of multiple human cancers. Herein, we report the preliminary anti-LSD1 evaluation of the synthetic vanadium (V) complexes. Among them, complex 2 showed a moderate inhibitory effect against LSD1 with IC₅₀ value of 19.0?μM, as well as good selectivity over MAO-A/B. Complex 2 is the first vanadium based LSD1 inhibitor, which provides a novel scaffold for the development of LSD1 inhibitor.     

Molecular modeling based approach, synthesis and in vitro assay to new indole inhibitors of hepatitis C NS3/4A serine protease

In an attempt to identify potential HCV NS3 protease inhibitors lead compounds, a series of novel indoles (10a-g) was designed. Molecular modeling study, including fitting to a 3D-pharmacophore model of the designed molecules (10a-g), with HCV NS3 protease hypothesis using catalyst program was fulfilled. Also, the molecular docking into the NS3 active site was examined using Discovery Studio 2.5 software. Several compounds showed significant high simulation docking score and fit values. The designed compounds with high docking score and fit values were synthesized and biologically evaluated in vitro using an NS3 protease binding assay. It appears that most of the tested compounds reveal promising inhibitory activity against NS3 protease. Of these, compounds 10a and 10b demonstrated potent HCV NS3 protease inhibitors with IC₅₀ values of 9 and 12 ??g/mL, respectively. The experimental serine protease inhibitor activities of compounds 10a-g were consistent with their molecular modeling results. Inhibitors from this class have promising characteristics for further development as anti-HCV agents.     

Baicalin,a natural LSD1 inhibitor

Baicalin is one of the active ingredients in the skullcap, with a variety of pharmacological effects, such as blood pressure reduction, sedation, liver-protection, gallbladder-protection, anti-bacteria, and anti-inflammation. In our study, baicalin was first characterized as a LSD1 inhibitor with an IC₅₀ of 3.01 μM and showed strong LSD1 inhibitory effect in cells. Baicalin may serve as a template for designing flavone-based LSD1 inhibitors.     

3D-QSAR (CoMFA,CoMSIA), molecular docking and molecular dynamics simulations study of 6-aryl-5-cyano-pyrimidine derivatives to explore the structure requirements of LSD1 inhibitors

Recently, Histone Lysine Specific Demethylase 1 (LSD1) was regarded as a promising anticancer target for the novel drug discovery. And several small molecules as LSD1 inhibitors in different structures have been reported. In this work, we carried out a molecular modeling study on the 6-aryl-5-cyano-pyrimidine fragment LSD1 inhibitors using three-dimensional quantitative structure–activity relationship (3D-QSAR), molecular docking and molecular dynamics simulations. Comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were used to generate 3D-QSAR models. The results show that the best CoMFA model has q² = 0.802, r²_ncv = 0.979, and the best CoMSIA model has q² = 0.799, r²_ncv = 0.982. The electrostatic, hydrophobic and H-bond donor fields play important roles in the models. Molecular docking studies predict the binding mode and the interactions between the ligand and the receptor protein. Molecular dynamics simulations results reveal that the complex of the ligand and the receptor protein are stable at 300 K. All the results can provide us more useful information for our further drug design.