Dataset Information

Peptide-Conjugated Phosphorodiamidate Morpholino Oligomers for In Situ Live-Cell Molecular Imaging of Dengue Virus Replication.

ABSTRACT: Current methods to detect and monitor pathogens in biological systems are largely limited by the tradeoffs between spatial context and temporal detail. A new generation of molecular tracking that provides both information simultaneously involves in situ detection coupled with non-invasive imaging. An example is antisense imaging that uses antisense oligonucleotide probes complementary to a target nucleotide sequence. In this study, we explored the potential of repurposing antisense oligonucleotides initially developed as antiviral therapeutics as molecular probes for imaging of viral infections in vitro and in vivo. We employed nuclease-resistant phosphorodiamidate synthetic oligonucleotides conjugated with cell-penetrating peptides (i.e., PPMOs) previously established as antivirals for dengue virus serotype-2 (DENV2). As proof of concept, and before further development for preclinical testing, we evaluated its validity as in situ molecular imaging probe for tracking cellular DENV2 infection using live-cell fluorescence imaging. Although the PPMO was designed to specifically target the DENV2 genome, it was unsuitable as in situ molecular imaging probe. This study details our evaluation of the PPMOs to assess specific and sensitive molecular imaging of DENV2 infection and tells a cautionary tale for those exploring antisense oligonucleotides as probes for non-invasive imaging and monitoring of pathogen infections in experimental animal models.

SUBMITTER: Victorio CBL

PROVIDER: S-EPMC7730579 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Peptide-Conjugated Phosphorodiamidate Morpholino Oligomers for In Situ Live-Cell Molecular Imaging of Dengue Virus Replication.

Victorio Carla Bianca Luena CBL Novera Wisna W Tham Jing Yang JY Watanabe Satoru S Vasudevan Subhash G SG Chacko Ann-Marie AM

International journal of molecular sciences 20201204 23

Current methods to detect and monitor pathogens in biological systems are largely limited by the tradeoffs between spatial context and temporal detail. A new generation of molecular tracking that provides both information simultaneously involves in situ detection coupled with non-invasive imaging. An example is antisense imaging that uses antisense oligonucleotide probes complementary to a target nucleotide sequence. In this study, we explored the potential of repurposing antisense oligonucleoti ...[more]

PMID: 33291644

Similar Datasets

Project description:In late 2015, the first example of a transferrable polymyxin resistance mechanism in Gram-negative pathogens, MCR-1, was reported. Since that report, MCR-1 has been described to occur in many Gram-negative pathogens, and the mechanism of MCR-1-mediated resistance was rapidly determined: an ethanolamine is attached to lipid A phosphate groups, rendering the membrane more electropositive and repelling positively charged polymyxins. Acquisition of MCR-1 is clinically significant because polymyxins are frequently last-line antibiotics used to treat extensively resistant organisms, so acquisition of this mechanism might lead to pan-resistant strains. Therefore, the ability to inhibit MCR-1 and restore polymyxin sensitivity would be an important scientific advancement. Peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs) are antisense molecules that were designed to target mRNA, preventing translation. Peptide conjugation enhances cellular entry, but they are positively charged, so we tested our lead antibacterial PPMOs by targeting an essential Escherichia coli gene, acpP, and demonstrated that they were still effective in mcr-1-positive E. coli strains. We then designed and synthesized two PPMOs targeted to mcr-1 mRNA. Five clinical mcr-1-positive E. coli strains were resensitized to polymyxins by MCR-1 inhibition, reducing MICs 2- to 16-fold. Finally, therapeutic dosing of BALB/c mice with MCR-1 PPMO combined with colistin in a sepsis model reduced morbidity and bacterial burden in the spleen at 24 h and offered a survival advantage out to 5 days. This is the first example of a way to modulate colistin resistance with an antisense approach and may be a viable strategy to combat this globally emerging antibiotic resistance threat.IMPORTANCE Polymyxin use has been increasing as a last line of defense against Gram-negative pathogens with high-level resistance mechanisms, such as carbapenemases. The recently described MCR-1 is a plasmid-mediated mechanism of resistance to polymyxins. MCR-1 is currently found in Gram-negative organisms already possessing high-level resistance mechanisms, leaving clinicians few or no antibacterial options for infections caused by these strains. This study utilizes antisense molecules that target mRNA, preventing protein translation. Herein we describe antisense molecules that can be directly antibacterial because they target genes essential to bacterial growth or blockade of MCR-1, restoring polymyxin sensitivity. We also demonstrate that MCR-1 antisense molecules restore the efficacies of polymyxins in mouse models of E. coli septicemia. Considering all things together, we demonstrate that antisense molecules may be effective therapeutics either alone when they target an essential gene or combined with antibiotics when they target specific resistance mechanisms, such as those seen with MCR-1.

Dataset Information

Peptide-Conjugated Phosphorodiamidate Morpholino Oligomers for In Situ Live-Cell Molecular Imaging of Dengue Virus Replication.

Publications

Peptide-Conjugated Phosphorodiamidate Morpholino Oligomers for In Situ Live-Cell Molecular Imaging of Dengue Virus Replication.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets