Brain-Penetrant LSD1 Inhibitors Can Block Memory Consolidation

Department of Radiology, Athinoula A. Martinos Center for Biomedical Imaging, Massachusetts General Hospital, Harvard Medical School, Charlestown, MA 02129, United States.
ACS Chemical Neuroscience (Impact Factor: 4.36). 02/2012; 3(2):120-128. DOI: 10.1021/cn200104y
Source: PubMed


Modulation of histone modifications in the brain may represent a new mechanism for brain disorder therapy. Post-translational modifications of histones regulate gene expression, affecting major cellular processes such as proliferation, differentiation, and function. An important enzyme involved in one of these histone modifications is lysine specific demethylase 1 (LSD1). This enzyme is flavin-dependent and exhibits homology to amine oxidases. Parnate (2-phenylcyclopropylamine (2-PCPA); tranylcypromine) is a potent inhibitor of monoamine oxidases and derivatives of 2-PCPA have been used for development of selective LSD1 inhibitors based on the ability to form covalent adducts with flavin adenine dinucleotide (FAD). Here we report the synthesis and in vitro characterization of LSD1 inhibitors that bond covalently to FAD. The two most potent and selective inhibitors were used to demonstrate brain penetration when administered systemically to rodents. First, radiosynthesis of a positron-emitting analog was used to obtain preliminary bio-distribution data and whole brain time-activity curves. Second, we demonstrate that this series of LSD1 inhibitors is capable of producing a cognitive effect in a mouse model. By using a memory formation paradigm, novel object recognition, we show that LSD1 inhibition can abolish long-term memory formation without affecting short-term memory, providing further evidence for the importance of reversible histone methylation in the function of the nervous system.

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    • "A series of patent applications were disclosed by Oryzon and GSK describing N-substituted tranylcypromine derivatives [41] [42], several of them being highly selective for KDM1A over MAO A and MAO B. Two compounds from these series have been reported to have entered clinical studies [43] [44]. Neelamegam et al. found that compound RN-7 (Fig. 1) was over 300 times more potent against KDM1A over MAO A and B [45]. The authors were able to demonstrate an involvement of KDM1A as regulator of long-term memory formation implying a key function in the central nervous system. "
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    ABSTRACT: Histone demethylase KDM1A (also known as LSD1) has become an attractive therapeutic target for the treatment of cancer as well as other disorders such as viral infections. We report on the synthesis of compounds derived from the expansion of tranylcypromine as a chemical scaffold for the design of novel demethylase inhibitors. These compounds, which are substituted on the cyclopropyl core moiety, were evaluated for their ability to inhibit KDM1A in vitro as well as to function in cells by modulating the expression of Gfi-1b, a well recognized KDM1A target gene. The molecules were all found to covalently inhibit KDM1A and to become increasingly selective against human monoamine oxidases MAO A and MAO B through the introduction of bulkier substituents on the cyclopropylamine ring. Structural and biochemical analysis of selected trans isomers showed that the two stereoisomers are endowed with similar inhibitory activities against KDM1A, but form different covalent adducts with the FAD co-enzyme.
    European Journal of Medicinal Chemistry 08/2014; 86C:352-363. DOI:10.1016/j.ejmech.2014.08.068 · 3.45 Impact Factor
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    • ") (Fig. 2A, Table 1), which are similar to reported results using same peptide substrate [35]. RN-1, an irreversible inhibitor of KDM1A [36], showed an IC 50 value of 0.6 ± 0.05 lM in our KDM1A/KMT7 coupled assay (Fig. 2B). We were able to perform this assay in a 384-well format with a Z 0 factor of 0.72 (Fig. S3) [37] "
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    ABSTRACT: Covalent modifications, such as methylation and demethylation of lysine residues in histones, play important roles in chromatin dynamics and the regulation of gene expression. The lysine demethylases (KDMs) catalyze the demethylation of lysine residues on histone tails and are associated with diverse human diseases, including cancer, and are therefore proposed as targets for the therapeutic modulation of gene transcription. High-throughput assays have been developed to find inhibitors of KDMs, most of which are fluorescence-based assays. Here we report the development of a coupled scintillation proximity assay (SPA) for 3 KDMs: KDM1A (LSD1), KDM3A (JMJD1A) and KDM4A (JMJD2A). In this assay methylated peptides are first demethylated by a KDM, and a protein methyltransferase (PMT) is added to methylate the resulting peptide with tritiated SAM. The enzyme activities were optimized and kinetic parameters were determined. These robust coupled assays are suitable for screening KDMs in 384-well format (Z'-Factors of 0.70-0.80) facilitating discovery of inhibitors in the quest for cancer therapeutics.
    Analytical Biochemistry 07/2014; 463. DOI:10.1016/j.ab.2014.06.023 · 2.22 Impact Factor
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    • "FAD-approved TCP has been used in the clinical treatment of mood and anxiety disorders [30-32]. Two of the most recently developed TCP-analogues with LSD1 inhibitory activity are S2101 [33] and RN-1 [34]. A third class of LSD1 inhibitors is based on FDA-approved antidepressants pargyline and phenelzine [25]. "
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    ABSTRACT: Lysine-specific demethylase 1 (LSD1, also known as KDM1A and AOF2) is a chromatin-modifying activity that catalyzes the removal of methyl groups from lysine residues in histone and non-histone proteins, regulating gene transcription. LSD1 is overexpressed in several cancer types, and chemical inhibition of the LSD1 activity has been proposed as a candidate cancer therapy. Here, we examine the levels of LSD1 mRNA in human ovarian tumors and the cytotoxicity of several chemical LSD1 inhibitors in a panel of ovarian cancer cell lines. We measured LSD1 mRNA levels in a cohort of n = 177 normal and heterogeneous tumor specimens by quantitative real time-PCR (qRT-PCR). Tumors were classified by FIGO stage, FIGO grade, and histological subtypes. We tested the robustness of our analyses in an independent cohort of n = 572 serous tumor specimens (source: TCGA, based on microarray). Statistical analyses were based on Kruskal-Wallis/Dunn's and Mann Whitney tests. Changes in LSD1 mRNA levels were also correlated with transcriptomic alterations at genome-wide scale. Effects on cell viability (MTS/PMS assay) of six LSD1 inhibitors (pargyline, TCP, RN-1, S2101, CAS 927019-63-4, and CBB1007) were also evaluated in a panel of ovarian cancer cell lines (SKOV3, OVCAR3, A2780 and cisplatin-resistant A2780cis). We found moderate but consistent LSD1 mRNA overexpression in stage IIIC and high-grade ovarian tumors. LSD1 mRNA overexpression correlated with a transcriptomic signature of up-regulated genes involved in cell cycle and down-regulated genes involved in the immune/inflammatory response, a signature previously observed in aggressive tumors. In fact, some ovarian tumors showing high levels of LSD1 mRNA are associated with poor patient survival. Chemical LSD1 inhibition induced cytotoxicity in ovarian cancer lines, which roughly correlated with their reported LSD1 inhibitory potential (RN-1,S2101> > pargyline,TCP). Our findings may suggest a role of LSD1 in the biology of some ovarian tumors. It is of special interest to find a correlation of LSD1 mRNA overexpression with a transcriptomic signature relevant to cancer. Our findings, therefore, prompt further investigation of the role of LSD1 in ovarian cancer, as well as the study of its enzymatic inhibition in animal models for potential therapeutic purposes in the context of this disease.
    Journal of Ovarian Research 10/2013; 6(1):75. DOI:10.1186/1757-2215-6-75 · 2.43 Impact Factor
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