Dataset Information

Selective Killing of Dormant Mycobacterium tuberculosis by Marine Natural Products.

ABSTRACT: The dormant phenotype acquired by Mycobacterium tuberculosis during infection poses a major challenge in disease treatment, since these bacilli show tolerance to front-line drugs. Therefore, it is imperative to find novel compounds that effectively kill dormant bacteria. By screening 4,400 marine natural product samples against dual-fluorescent M. tuberculosis under both replicating and nonreplicating conditions, we have identified compounds that are selectively active against dormant M. tuberculosis This validates our strategy of screening all compounds in both assays as opposed to using the dormancy model as a secondary screen. Bioassay-guided deconvolution enabled the identification of unique pharmacophores active in each screening model. To confirm the activity of samples against dormant M. tuberculosis, we used a luciferase reporter assay and enumerated CFU. The structures of five purified active compounds were defined by nuclear magnetic resonance (NMR) and mass spectrometry. We identified two lipid compounds with potent activity toward dormant and actively growing M. tuberculosis strains. One of these was commercially obtained and showed similar activity against M. tuberculosis in both screening models. Furthermore, puupehenone-like molecules were purified with potent and selective activity against dormant M. tuberculosis In conclusion, we have identified and characterized antimycobacterial compounds from marine organisms with novel activity profiles which appear to target M. tuberculosis pathways that are conditionally essential for dormancy survival.

SUBMITTER: Rodrigues Felix C

PROVIDER: S-EPMC5527660 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Selective Killing of Dormant Mycobacterium tuberculosis by Marine Natural Products.

Rodrigues Felix Carolina C Gupta Rashmi R Geden Sandra S Roberts Jill J Winder Priscilla P Pomponi Shirley A SA Diaz Maria Cristina MC Reed John K JK Wright Amy E AE Rohde Kyle H KH

Antimicrobial agents and chemotherapy 20170725 8

The dormant phenotype acquired by <i>Mycobacterium tuberculosis</i> during infection poses a major challenge in disease treatment, since these bacilli show tolerance to front-line drugs. Therefore, it is imperative to find novel compounds that effectively kill dormant bacteria. By screening 4,400 marine natural product samples against dual-fluorescent <i>M. tuberculosis</i> under both replicating and nonreplicating conditions, we have identified compounds that are selectively active against dorm ...[more]

PMID: 28607021

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Project description:Tuberculosis is a disease caused primarily by the organism Mycobacterium tuberculosis (Mtb), which claims about 1.5 million lives every year. A challenge that impedes the elimination of this pathogen is the ability of Mtb to remain dormant after primary infection, thus creating a reservoir for the disease in the population that reactivates under more ideal conditions. A better understanding of the physiology of dormant Mtb and therapeutics able to kill these phenotypically tolerant bacilli will be critical for completely eradicating Mtb. Our groups are focusing on characterizing the activity of derivatives of the marine natural product (+)-puupehenone (1). Recently, the Rohde group reported that puupehedione (2) and 15-α-methoxypuupehenol (3) exhibit enhanced activity in an in vitro multi-stress dormancy model of Mtb. To optimize the antimycobacterial activity of these terpenoids, novel 15-α-methoxy- and 15-α-acetoxy-puupehenol esters were prepared from (+)-puupehenone (1) accessed through a (+)-sclareolide-derived β-hydroxyl aldehyde. For added diversity, various congeners related to (1) were also prepared from a common borono-sclareolide donor, which resulted in the synthesis of epi-puupehenol and the natural products (+)-chromazonarol and (+)-yahazunol. In total, we generated a library of 24 compounds, of which 14 were found to be active against Mtb, and the most active compounds retained the enhanced activity against dormant Mtb seen in the parent compound. Several of the 15-α-methoxy- and 15-α-acetoxy-puupehenol esters possessed potent activity against actively dividing and dormant Mtb. Intriguingly, the closely related triisobutyl derivative 16 showed similar activity to 1 in actively dividing Mtb but lost about 178-fold activity against dormant Mtb. However, the monopivaloyl compound 13 showed a modest 3- to 4-fold loss in activity in both actively dividing and dormant Mtb relative to the activity of 1 revealing the importance of the free OH at C19 supporting the potential role of quinone methide formation as critical for activity in dormant Mtb. Elucidating important structure-activity relationships and the mechanism of action of this natural product-inspired chemical series may yield insights into vulnerable drug targets in dormant bacilli and new therapeutics to more effectively target dormant Mtb.

Project description:Dormant Mycobacterium tuberculosis bacilli are believed to play an important role in latent tuberculosis infection. Previously, we have demonstrated that cultivation of M. tuberculosis in K(+)-deficient medium resulted in generation of dormant cells. These bacilli were non-culturable on solid media (a key feature of dormant M. tuberculosis in vivo) and characterized by low metabolism and tolerance to anti-tuberculosis drugs. The dormant bacteria demonstrated a high potential to reactivation after K(+) reintroduction even after prolonged persistence under rifampicin. In this work, we studied the transcriptome and stability of transcripts in persisting dormant bacilli under arrest of mRNA de novo synthesis.RNA-seq-based analysis of the dormant non-culturable population obtained under rifampicin exposure revealed a 30-50-fold decrease of the total mRNA level, indicating global transcriptional repression. However, the analysis of persisting transcripts displayed a cohort of mRNA molecules coding for biosynthetic enzymes, proteins involved in adaptation and repair processes, detoxification, and control of transcription initiation. This 'dormant transcriptome' demonstrated considerable stability during M. tuberculosis persistence and mRNA de novo synthesis arrest. On the contrary, several small non-coding RNAs showed increased abundance on dormancy. Interestingly, M. tuberculosis entry into dormancy was accompanied by the cleavage of 23S ribosomal RNA at a specific point located outside the ribosome catalytic center.Dormant non-culturable M. tuberculosis bacilli are characterized by a global transcriptional repression. At the same time, the dormant bacilli retain low-abundant mRNAs, which are considerably stable during in vitro persistence, reflecting their readiness for translation upon early resuscitation steps. Increased abundance of non-coding RNAs on dormancy may indicate their role in the entry into and maintenance of M. tuberculosis dormant non-culturable state.

Dataset Information

Selective Killing of Dormant Mycobacterium tuberculosis by Marine Natural Products.

Publications

Selective Killing of Dormant Mycobacterium tuberculosis by Marine Natural Products.

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