Characterization and structure-activity relationships of indenoisoquinoline-derived topoisomerase I inhibitors in unsilencing the dormant Ube3a gene associated with Angelman syndrome / Hyeong-Min LEE in Molecular Autism, 9 (2018)  

Public ISBD [article]  inMolecular Autism > 9 (2018) . - 45p. Titre : Characterization and structure-activity relationships of indenoisoquinoline-derived topoisomerase I inhibitors in unsilencing the dormant Ube3a gene associated with Angelman syndrome Type de document : texte imprimé Auteurs : Hyeong-Min LEE, Auteur ; Ellen P. CLARK, Auteur ; M. Bram KUIJER, Auteur ; Mark CUSHMAN, Auteur ; Yves POMMIER, Auteur ; Benjamin D. PHILPOT, Auteur Article en page(s) : 45p. Langues : Anglais (eng) Mots-clés : Angelman syndrome  Indenoisoquinoline  Indotecan  Topoisomerase I  Topoisomerase inhibitor  Topotecan  UBE3A Index. décimale : PER Périodiques Résumé : Background: Angelman syndrome (AS) is a severe neurodevelopmental disorder lacking effective therapies. AS is caused by mutations in ubiquitin protein ligase E3A (UBE3A), which is genomically imprinted such that only the maternally inherited copy is expressed in neurons. We previously demonstrated that topoisomerase I (Top1) inhibitors could successfully reactivate the dormant paternal allele of Ube3a in neurons of a mouse model of AS. We also previously showed that one such Top1 inhibitor, topotecan, could unsilence paternal UBE3A in induced pluripotent stem cell-derived neurons from individuals with AS. Although topotecan has been well-studied and is FDA-approved for cancer therapy, its limited CNS bioavailability will likely restrict the therapeutic use of topotecan in AS. The goal of this study was to identify additional Top1 inhibitors with similar efficacy as topotecan, with the expectation that these could be tested in the future for safety and CNS bioavailability to assess their potential as AS therapeutics. Methods: We tested 13 indenoisoquinoline-derived Top1 inhibitors to identify compounds that unsilence the paternal allele of Ube3a in mouse neurons. Primary cortical neurons were isolated from embryonic day 14.5 (E14.5) mice with a Ube3a-YFP fluorescent tag on the paternal allele (Ube3a(m+/pYFP) mice) or mice that lack the maternal Ube3a allele and hence model AS (Ube3a(m-/p+) mice). Neurons were cultured for 7 days, treated with drug for 72 h, and examined for paternal UBE3A protein expression by Western blot or fluorescence immunostaining. Dose responses of the compounds were determined across a log range of drug treatments, and cytotoxicity was tested using a luciferase-based assay. Results: All 13 indenoisoquinoline-derived Top1 inhibitors unsilenced paternal Ube3a. Several compounds exhibited favorable paternal Ube3a unsilencing properties, similar to topotecan, and of these, indotecan (LMP400) was the most effective based on estimated Emax (maximum response of unsilencing paternal Ube3a) and EC50 (half maximal effective concentration). Conclusions: We provide pharmacological profiles of indenoisoquinoline-derived Top1 inhibitors as paternal Ube3a unsilencers. All 13 tested compounds were effective at unsilencing paternal Ube3a, although with variable efficacy and potency. Indotecan (LMP400) demonstrated a better pharmacological profile of Ube3a unsilencing compared to our previous lead compound, topotecan. Taken together, indotecan and its structural analogues are potential AS therapeutics whose translational potential in AS treatment should be further assessed. En ligne : https://dx.doi.org/10.1186/s13229-018-0228-2 Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=371 [article] Characterization and structure-activity relationships of indenoisoquinoline-derived topoisomerase I inhibitors in unsilencing the dormant Ube3a gene associated with Angelman syndrome [texte imprimé] / Hyeong-Min LEE, Auteur ; Ellen P. CLARK, Auteur ; M. Bram KUIJER, Auteur ; Mark CUSHMAN, Auteur ; Yves POMMIER, Auteur ; Benjamin D. PHILPOT, Auteur . - 45p. Langues : Anglais (eng) in Molecular Autism > 9 (2018) . - 45p. Mots-clés : Angelman syndrome  Indenoisoquinoline  Indotecan  Topoisomerase I  Topoisomerase inhibitor  Topotecan  UBE3A Index. décimale : PER Périodiques Résumé : Background: Angelman syndrome (AS) is a severe neurodevelopmental disorder lacking effective therapies. AS is caused by mutations in ubiquitin protein ligase E3A (UBE3A), which is genomically imprinted such that only the maternally inherited copy is expressed in neurons. We previously demonstrated that topoisomerase I (Top1) inhibitors could successfully reactivate the dormant paternal allele of Ube3a in neurons of a mouse model of AS. We also previously showed that one such Top1 inhibitor, topotecan, could unsilence paternal UBE3A in induced pluripotent stem cell-derived neurons from individuals with AS. Although topotecan has been well-studied and is FDA-approved for cancer therapy, its limited CNS bioavailability will likely restrict the therapeutic use of topotecan in AS. The goal of this study was to identify additional Top1 inhibitors with similar efficacy as topotecan, with the expectation that these could be tested in the future for safety and CNS bioavailability to assess their potential as AS therapeutics. Methods: We tested 13 indenoisoquinoline-derived Top1 inhibitors to identify compounds that unsilence the paternal allele of Ube3a in mouse neurons. Primary cortical neurons were isolated from embryonic day 14.5 (E14.5) mice with a Ube3a-YFP fluorescent tag on the paternal allele (Ube3a(m+/pYFP) mice) or mice that lack the maternal Ube3a allele and hence model AS (Ube3a(m-/p+) mice). Neurons were cultured for 7 days, treated with drug for 72 h, and examined for paternal UBE3A protein expression by Western blot or fluorescence immunostaining. Dose responses of the compounds were determined across a log range of drug treatments, and cytotoxicity was tested using a luciferase-based assay. Results: All 13 indenoisoquinoline-derived Top1 inhibitors unsilenced paternal Ube3a. Several compounds exhibited favorable paternal Ube3a unsilencing properties, similar to topotecan, and of these, indotecan (LMP400) was the most effective based on estimated Emax (maximum response of unsilencing paternal Ube3a) and EC50 (half maximal effective concentration). Conclusions: We provide pharmacological profiles of indenoisoquinoline-derived Top1 inhibitors as paternal Ube3a unsilencers. All 13 tested compounds were effective at unsilencing paternal Ube3a, although with variable efficacy and potency. Indotecan (LMP400) demonstrated a better pharmacological profile of Ube3a unsilencing compared to our previous lead compound, topotecan. Taken together, indotecan and its structural analogues are potential AS therapeutics whose translational potential in AS treatment should be further assessed. En ligne : https://dx.doi.org/10.1186/s13229-018-0228-2 Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=371

A small-molecule screen reveals novel modulators of MeCP2 and X-chromosome inactivation maintenance / Hyeong-Min LEE in Journal of Neurodevelopmental Disorders, 12 (2020)  

Public ISBD [article]  inJournal of Neurodevelopmental Disorders > 12 (2020) Titre : A small-molecule screen reveals novel modulators of MeCP2 and X-chromosome inactivation maintenance Type de document : texte imprimé Auteurs : Hyeong-Min LEE, Auteur ; M Bram KUIJER, Auteur ; Nerea RUIZ BLANES, Auteur ; Ellen P. CLARK, Auteur ; Megumi AITA, Auteur ; Lorena GALIANO ARJONA, Auteur ; Agnieszka KOKOT, Auteur ; Noah SCIAKY, Auteur ; Jeremy M. SIMON, Auteur ; Sanchita BHATNAGAR, Auteur ; Benjamin D. PHILPOT, Auteur ; Andrea CERASE, Auteur Langues : Anglais (eng) Mots-clés : Animals  Chromosomes  Female  Male  Methyl-CpG-Binding Protein 2/genetics/metabolism  Mice  Mutation  Rett Syndrome/drug therapy/genetics  X Chromosome Inactivation  Ag490  Jak/stat  Janus Kinase  Janus Kinase inhibitors  MeCP2  PI3K/ATK pathways  Rett syndrome  X-chromosome inactivation Index. décimale : PER Périodiques Résumé : BACKGROUND: Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X-linked methyl-CpG binding protein 2 (MeCP2) gene. While MeCP2 mutations are lethal in most males, females survive birth but show severe neurological defects. Because X-chromosome inactivation (XCI) is a random process, approximately 50% of the cells silence the wild-type (WT) copy of the MeCP2 gene. Thus, reactivating the silent WT copy of MeCP2 could provide therapeutic intervention for RTT. METHODS: Toward this goal, we screened ~ 28,000 small-molecule compounds from several libraries using a MeCP2-luciferase reporter cell line and cortical neurons from a MeCP2-EGFP mouse model. We used gain/increase of luminescence or fluorescence as a readout of MeCP2 reactivation and tested the efficacy of these drugs under different drug regimens, conditions, and cellular contexts. RESULTS: We identified inhibitors of the JAK/STAT pathway as XCI-reactivating agents, both by in vitro and ex vivo assays. In particular, we show that AG-490, a Janus Kinase 2 (JAK2) kinase inhibitor, and Jaki, a pan JAK/STAT inhibitor, are capable of reactivating MeCP2 from the inactive X chromosome, in different cellular contexts. CONCLUSIONS: Our results suggest that inhibition of the JAK/STAT pathway is a new potential pathway to reinstate MeCP2 gene expression as an efficient RTT treatment. En ligne : https://dx.doi.org/10.1186/s11689-020-09332-3 Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=573 [article] A small-molecule screen reveals novel modulators of MeCP2 and X-chromosome inactivation maintenance [texte imprimé] / Hyeong-Min LEE, Auteur ; M Bram KUIJER, Auteur ; Nerea RUIZ BLANES, Auteur ; Ellen P. CLARK, Auteur ; Megumi AITA, Auteur ; Lorena GALIANO ARJONA, Auteur ; Agnieszka KOKOT, Auteur ; Noah SCIAKY, Auteur ; Jeremy M. SIMON, Auteur ; Sanchita BHATNAGAR, Auteur ; Benjamin D. PHILPOT, Auteur ; Andrea CERASE, Auteur. Langues : Anglais (eng) in Journal of Neurodevelopmental Disorders > 12 (2020) Mots-clés : Animals  Chromosomes  Female  Male  Methyl-CpG-Binding Protein 2/genetics/metabolism  Mice  Mutation  Rett Syndrome/drug therapy/genetics  X Chromosome Inactivation  Ag490  Jak/stat  Janus Kinase  Janus Kinase inhibitors  MeCP2  PI3K/ATK pathways  Rett syndrome  X-chromosome inactivation Index. décimale : PER Périodiques Résumé : BACKGROUND: Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X-linked methyl-CpG binding protein 2 (MeCP2) gene. While MeCP2 mutations are lethal in most males, females survive birth but show severe neurological defects. Because X-chromosome inactivation (XCI) is a random process, approximately 50% of the cells silence the wild-type (WT) copy of the MeCP2 gene. Thus, reactivating the silent WT copy of MeCP2 could provide therapeutic intervention for RTT. METHODS: Toward this goal, we screened ~ 28,000 small-molecule compounds from several libraries using a MeCP2-luciferase reporter cell line and cortical neurons from a MeCP2-EGFP mouse model. We used gain/increase of luminescence or fluorescence as a readout of MeCP2 reactivation and tested the efficacy of these drugs under different drug regimens, conditions, and cellular contexts. RESULTS: We identified inhibitors of the JAK/STAT pathway as XCI-reactivating agents, both by in vitro and ex vivo assays. In particular, we show that AG-490, a Janus Kinase 2 (JAK2) kinase inhibitor, and Jaki, a pan JAK/STAT inhibitor, are capable of reactivating MeCP2 from the inactive X chromosome, in different cellular contexts. CONCLUSIONS: Our results suggest that inhibition of the JAK/STAT pathway is a new potential pathway to reinstate MeCP2 gene expression as an efficient RTT treatment. En ligne : https://dx.doi.org/10.1186/s11689-020-09332-3 Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=573

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