title: High-content screening of natural products reveals novel nuclear export inhibitors.
abstract: Natural products are considered an extremely valuable source for the discovery of new drugs against diverse pathologies. As yet, we have only explored a fraction of the diversity of bioactive compounds, and opportunities for discovering new natural products leading to new drugs are huge. In the present study, U2nesRELOC, a previously established cell-based imaging assay, was employed to screen a collection of extracts of microbial origin for nuclear export inhibition activity. The fluorescent signal of untreated U2nesRELOC cells localizes predominantly to the cytoplasm. Upon treatment with the nuclear export inhibitor leptomycin B, the fluorescent-tagged reporter proteins appear as speckles in the nucleus. A proprietary collection of extracts from fungi, actinomycetes, and unicellular bacteria that covers an uncommonly broad chemical space was used to interrogate this nuclear export assay system. A two-step image-based analysis allowed us to identify 12 extracts with biological activities that are not associated with previously known active metabolites. The fractionation and structural elucidation of active compounds revealed several chemical structures with nuclear export inhibition activity. Here we show that substrates of the nuclear export receptor CRM1, such as Rev, FOXO3a and NF-κB, accumulate in the nucleus in the presence of the fungal metabolite MDN-0105 with an IC50 value of 3.4 µM. Many important processes in tumor formation and progression, as well as in many viral infections, critically depend on the nucleocytoplasmic trafficking of proteins and RNA molecules. Therefore, the disruption of nuclear export is emerging as a novel therapeutic approach with enormous clinical potential. Our work highlights the potential of applying high-throughput phenotypic imaging on natural product extracts to identify novel nuclear export inhibitors.
authors.affiliation: 1Fundación MEDINA, Parque Tecnológico Ciencias de la Salud, Granada, España.
authors.forename: Bastien
authors.initials: B
authors.lastname: Cautain
authors.forename: Nuria
authors.initials: N
authors.lastname: de Pedro
authors.forename: Virginia
authors.initials: V
authors.lastname: Murillo Garzón
authors.forename: María
authors.initials: M
authors.lastname: Muñoz de Escalona
authors.forename: Victor
authors.initials: V
authors.lastname: González Menéndez
authors.forename: José R
authors.initials: JR
authors.lastname: Tormo
authors.forename: Jesús
authors.initials: J
authors.lastname: Martin
authors.forename: Noureddine
authors.initials: N
authors.lastname: El Aouad
authors.forename: Fernando
authors.initials: F
authors.lastname: Reyes
authors.forename: Francisco
authors.initials: F
authors.lastname: Asensio
authors.forename: Olga
authors.initials: O
authors.lastname: Genilloud
authors.forename: Francisca
authors.initials: F
authors.lastname: Vicente
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Link
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects*; D000818:Animals; D001688:Biological Products / Q000302:isolation & purification / Q000494:pharmacology*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D005591:Chemical Fractionation / Q000379:methods; D004305:Dose-Response Relationship, Drug; D055808:Drug Discovery / Q000379:methods; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D051858:Forkhead Transcription Factors / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D017930:Genes, Reporter; D057166:High-Throughput Screening Assays; D006801:Humans; D020128:Inhibitory Concentration 50; D016328:NF-kappa B / Q000378:metabolism
chemical_list: D001688:Biological Products; D051858:Forkhead Transcription Factors; D016328:NF-kappa B
keywords: high-content screening; natural products screening; oncology; phenotypic drug discovery; reporter gene assays