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Observing an Antisense Drug Complex in Intact Human Cells by in-Cell NMR Spectroscopy
Stockholm University, Faculty of Science, Department of Materials and Environmental Chemistry (MMK).
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Number of Authors: 92019 (English)In: ChemBioChem (Print), ISSN 1439-4227, E-ISSN 1439-7633, Vol. 20, p. 2474-2478Article in journal (Refereed) Published
Abstract [en]

Gaining insight into the uptake, trafficking and target engagement of drugs in cells can enhance understanding of a drug's function and efficiency. However, there are currently no reliable methods for studying untagged biomolecules in macromolecular complexes in intact human cells. Here we have studied an antisense oligonucleotide (ASO) drug in HEK 293T and HeLa cells by NMR spectroscopy. Using a combination of transfection, cryoprotection and dynamic nuclear polarization (DNP), we were able to detect the drug directly in intact frozen cells. Activity of the drug was confirmed by quantitative reverse transcription polymerase chain reaction (qRT-PCR). By applying DNP NMR to frozen cells, we overcame limitations both of solution-state in-cell NMR spectroscopy (e.g., size, stability and sensitivity) and of visualization techniques, in which (e.g., fluorescent) tagging of the ASO decreases its activity. The capability to detect an untagged, active drug, interacting in its natural environment, represents a first step towards studying molecular mechanisms in intact cells.

Place, publisher, year, edition, pages
2019. Vol. 20, p. 2474-2478
Keywords [en]
antisense agents, dynamic nuclear polarization, in-cell NMR spectroscopy, nucleic acids, oligonucleotides
National Category
Chemical Sciences
Identifiers
URN: urn:nbn:se:su:diva-175080DOI: 10.1002/cbic.201900297ISI: 000486747500001PubMedID: 31206961OAI: oai:DiVA.org:su-175080DiVA, id: diva2:1367662
Available from: 2019-11-04 Created: 2019-11-04 Last updated: 2019-11-04Bibliographically approved

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Jaworski, AleksanderPell, Andrew J.
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