Journal
DRUG METABOLISM AND DISPOSITION
Volume 43, Issue 7, Pages 1129-1136Publisher
AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS
DOI: 10.1124/dmd.115.064493
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Funding
- National Institutes of Health National Cancer Institute [1U01CA175315]
- National Science Foundation [CHE 1212625]
- Natural Science Foundation of China [81320108027]
- Ministry of Science and Technology of China [2012ZX09506001-004]
- Direct For Mathematical & Physical Scien
- Division Of Chemistry [1212625] Funding Source: National Science Foundation
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In contrast to the growing interests in studying noncoding RNAs (ncRNAs) such as microRNA (miRNA or miR) pharmacoepigenetics, there is a lack of efficient means to cost effectively produce large quantities of natural miRNA agents. Our recent efforts led to a successful production of chimeric pre-miR-27b in bacteria using a transfer RNA (tRNA)-based recombinant RNA technology, but at very low expression levels. Herein, we present a high-yield expression of chimeric pre-miR-1291 in common Escherichia coli strains using the same tRNA scaffold. The tRNA fusion pre-miR1291 (tRNA/mir-1291) was then purified to high homogeneity using affinity chromatography, whose primary sequence and posttranscriptional modifications were directly characterized by mass spectrometric analyses. Chimeric tRNA/mir-1291 was readily processed to mature miR-1291 in human carcinoma MCF-7 and PANC-1 cells. Consequently, recombinant tRNA/mir-1291 reduced the protein levels of miR-1291 target genes, including ABCC1, FOXA2, and MeCP2, as compared with cells transfected with the same doses of control methionyl-tRNA scaffold with a sephadex aptamer (tRNA/MSA). In addition, tRNA-carried premiR-1291 suppressed the growth of MCF-7 and PANC-1 cells in a dose-dependent manner, and significantly enhanced the sensitivity of ABCC1-overexpressing PANC-1 cells to doxorubicin. These results indicate that recombinant miR-1291 agent is effective in the modulation of target gene expression and chemosensitivity, which may provide insights into high-yield bioengineering of new ncRNA agents for pharmacoepigenetics research.
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